Based on a combination of next-generation sequencing and single-molecule sequencing, we obtained the whole-genome sequence of Bacillus megaterium strain TG1-E1, which is a highly salt-tolerant rhizobacterium that enhances plant tolerance to drought stress. The complete genome is estimated to be approximately 5.48 Mb containing a total of 5,858 predicted protein-coding DNA sequences.
ABSTRACT
Based on a combination of next-generation sequencing and single-molecule sequencing, we obtained the whole-genome sequence of Bacillus megaterium strain TG1-E1, which is a highly salt-tolerant rhizobacterium that enhances plant tolerance to drought stress. The complete genome is estimated to be approximately 5.48 Mb containing a total of 5,858 predicted protein-coding DNA sequences.
ANNOUNCEMENT
Our group characterized Bacillus megaterium strain TG1-E1 as a highly salt-tolerant Gram-positive bacterium that is capable of enhancing plant tolerance to drought stress. It was originally isolated from a rhizospheric soil sample of Spartina anglica at Zhangpu Yanchang in Fujian Province, China. This rhizobacterium collection is rich in specimens of the Firmicutes and Proteobacteria phyla, with about 70% belonging to the Bacillaceae family. High salinity in the sampling area possibly contributes to the enrichment of Bacillus strains in the rhizosphere (1–4). In addition, more than half of the strains isolated in this sampling area can produce phytometabolites, such as auxins and aminocyclopropane-1-carboxylate deaminase (ACCd), displaying the characteristics commonly described in plant tolerance-enhancing strains (5–10). B. megaterium TG1-E1 has been deposited in the China General Microbiological Culture Collection Center (CGMCC) with reference number 14422.
DNA samples (at least 100 nM in 10 µl) were obtained from bacteria grown in LB medium until an optical density of 1 at 600 nm (OD600) was obtained. The sequencing of the B. megaterium TG1-E1 genome was completed by combining next-generation sequencing (NGS) and single-molecule sequencing. NGS was performed with 20 µg of DNA with an Illumina HiSeq platform (Core Facility of Genomics, Shanghai Center for Plant Stress Biology, China), and single-molecule sequencing was performed with 20 µg of DNA with a PacBio platform (Tianjin Biochip Corporation, China) (11–14). The shotgun sequencing strategy was applied to NGS, and 12,471,203 paired reads (150 bp) were obtained with a sequencing depth of approximately 260-fold. Meanwhile, single-molecule sequencing produced 98,959 reads with a mean read length of 10,551 bp and an N50 length of 14,471 bp. The total number of sequenced bases was 961,774,920. For de novo assembly, Canu v1.5 was used with default parameters, and the genome correction step was performed using Illumina data with support of Pilon v1.18 (15, 16). The size of the circularized genome was calculated to be about 5.48 Mb. Genes including protein-coding DNA sequences (CDSs) were predicted by a pipeline implemented by Prokka v1.12 (17). On a whole-genome scale, The GC content of this genome is 38.26%, and 5,858 protein-coding genes, 4 rRNA operons, and 164 tRNA genes were called during annotation.
The whole-genome sequence of B. megaterium TG1-E1 reveals information such as the biosynthesis pathways of flagella, spores, and polysaccharides. Concerning characteristics potentially contributing to TG1-E1-induced plant stress tolerance, pathways found within this genome that have potential relevance in aiding plant drought stress include trehalose and antioxidant biosynthesis. In addition, genome annotation also revealed possible mechanisms for plant growth-promoting effects, including bacterial production of acid phosphatases, siderophores, and exopolysaccharides. Further research with this genomic information will help us discover mechanisms through which B. megaterium TG1-E1 induces plant drought stress tolerance and will contribute to the subsequent development of biotechnological applications.
Data availability.
The complete genome sequence of B. megaterium TG1-E1 has been deposited in the TBL/EMBL/GenBank databases under the BioProject number PRJNA430758 and the accession number PRKV00000000 (sequences PRKV01000001 to PRKV01000036).
ACKNOWLEDGMENTS
Huiming Zhang is funded by the Chinese Academy of Sciences (CAS) project number XDPB0404.
We acknowledge the support of the Core Facility of Genomics at PSC.
REFERENCES
- 1.Govindasamy V, Senthilkumar M, Magheshwaran V, Kumar U, Bose P, Sharma V, Annapurna K, 2011. Bacillus and Paenibacillus spp.: potential PGPR for sustainable agriculture, p 333–364. In Maheshwari DK. (ed), Plant growth and health promoting bacteria. Springer, Berlin, Germany. doi: 10.1007/978-3-642-13612-2_15. [DOI] [Google Scholar]
- 2.Nautiyal CS, Srivastava S, Chauhan PS, Seem K, Mishra A, Sopory SK. 2013. Plant growth-promoting bacteria Bacillus amyloliquefaciens NBRISN13 modulates gene expression profile of leaf and rhizosphere community in rice during salt stress. Plant Physiol Biochem 66:1–9. doi: 10.1016/j.plaphy.2013.01.020. [DOI] [PubMed] [Google Scholar]
- 3.Orhan F, Gulluce M. 2015. Isolation and characterization of salt-tolerant bacterial strains in salt-affected soils of east Anatolian region. Geomicrobiol J 32:10–16. doi: 10.1080/01490451.2014.917743. [DOI] [Google Scholar]
- 4.Yaish MW, Al-Lawati A, Jana GA, Vishwas Patankar H, Glick BR. 2016. Impact of soil salinity on the structure of the bacterial endophytic community identified from the roots of Caliph medic (Medicago truncatula). PLoS One 11:e0159007. doi: 10.1371/journal.pone.0159007. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Vurukonda SSKP, Vardharajula S, Shrivastava M, SkZ A. 2016. Enhancement of drought stress tolerance in crops by plant growth promoting rhizobacteria. Microbiol Res 184:13–24. doi: 10.1016/j.micres.2015.12.003. [DOI] [PubMed] [Google Scholar]
- 6.Vílchez JI, García-Fontana C, Román-Naranjo D, González-López J, Manzanera M. 2016. Plant drought tolerance enhancement by trehalose production of desiccation-tolerant microorganisms. Front Microbiol 7:1577. doi: 10.3389/fmicb.2016.01577. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.de Souza R, Ambrosini A, Passaglia LMP. 2015. Plant growth-promoting bacteria as inoculants in agricultural soils. Genet Mol Biol 38:401–419. doi: 10.1590/S1415-475738420150053. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Glick BR. 2012. Plant growth-promoting bacteria: mechanisms and applications. Scientifica 2012:1. doi: 10.6064/2012/963401. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.Kang S-M, Radhakrishnan R, You Y-H, Joo G-J, Lee I-J, Lee K-E, Kim J-H. 2014. Phosphate solubilizing Bacillus megaterium mj1212 regulates endogenous plant carbohydrates and amino acids contents to promote mustard plant growth. Indian J Microbiol 54:427–433. doi: 10.1007/s12088-014-0476-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Santos S, Neto IF, Machado MD, Soares HM, Soares EV. 2014. Siderophore production by Bacillus megaterium: effect of growth phase and cultural conditions. Appl Biochem Biotechnol 172:549–560. doi: 10.1007/s12010-013-0562-y. [DOI] [PubMed] [Google Scholar]
- 11.Rhoads A, Au KF. 2015. PacBio sequencing and its applications. Genomics Proteomics Bioinformatics 13:278–289. doi: 10.1016/j.gpb.2015.08.002. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Eid J, Fehr A, Gray J, Luong K, Lyle J, Otto G, Peluso P, Rank D, Baybayan P, Bettman B, Bibillo A, Bjornson K, Chaudhuri B, Christians F, Cicero R, Clark S, Dalal R, Dewinter A, Dixon J, Foquet M, Gaertner A, Hardenbol P, Heiner C, Hester K, Holden D, Kearns G, Kong X, Kuse R, Lacroix Y, Lin S, Lundquist P, Ma C, Marks P, Maxham M, Murphy D, Park I, Pham T, Phillips M, Roy J, Sebra R, Shen G, Sorenson J, Tomaney A, Travers K, Trulson M, Vieceli J, Wegener J, Wu D, Yang A, Zaccarin D, Zhao P, Zhong F, Korlach J, Turner S. 2009. Real-time DNA sequencing from single polymerase molecules. Science 323:133–138. doi: 10.1126/science.1162986. [DOI] [PubMed] [Google Scholar]
- 13.Buermans HPJ, den Dunnen JT. 2014. Next generation sequencing technology: advances and applications. Biochim Biophys Acta 1842:1932–1941. doi: 10.1016/j.bbadis.2014.06.015. [DOI] [PubMed] [Google Scholar]
- 14.Huptas C, Scherer S, Wenning M. 2016. Optimized Illumina PCR-free library preparation for bacterial whole genome sequencing and analysis of factors influencing de novo assembly. BMC Res Notes 9:269. doi: 10.1186/s13104-016-2072-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Walker BJ, Abeel T, Shea T, Priest M, Abouelliel A, Sakthikumar S, Cuomo CA, Zeng Q, Wortman J, Young SK, Earl AM. 2014. Pilon: an integrated tool for comprehensive microbial variant detection and genome assembly improvement. PLoS One 9:e112963. doi: 10.1371/journal.pone.0112963. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Koren S, Walenz BP, Berlin K, Miller JR, Bergman NH, Phillippy AM. 2017. Canu: scalable and accurate long-read assembly via adaptive k-mer weighting and repeat separation. Genome Res 27:722–736. doi: 10.1101/gr.215087.116. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17.Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bioinformatics 30:2068–2069. doi: 10.1093/bioinformatics/btu153. [DOI] [PubMed] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
The complete genome sequence of B. megaterium TG1-E1 has been deposited in the TBL/EMBL/GenBank databases under the BioProject number PRJNA430758 and the accession number PRKV00000000 (sequences PRKV01000001 to PRKV01000036).