1887

Abstract

Ensifer meliloti (formerly Rhizobium meliloti and Sinorhizobium meliloti) is a model bacterium for understanding legume–rhizobial symbioses. The tripartite genome of E. meliloti consists of a chromosome, pSymA and pSymB, and in some instances strain-specific accessory plasmids. The majority of previous sequencing studies have relied on the use of assemblies generated from short read sequencing, which leads to gaps and assembly errors. Here we used PacBio-based, long-read assemblies and were able to assemble, de novo, complete circular replicons. In this study, we sequenced, de novo-assembled and analysed 10 E. meliloti strains. Sequence comparisons were also done with data from six previously published genomes. We identified genome differences between the replicons, including mol% G+C and gene content, nucleotide repeats, and transposon-associated loci. Additionally, genomic rearrangements both within and between replicons were identified, providing insight into evolutionary processes at the structural level. There were few cases of inter-replicon gene transfer of core genes between the main replicons. Accessory plasmids were more similar to pSymA than to either pSymB or the chromosome, with respect to gene content, transposon content and G+C content. In our population, the accessory plasmids appeared to share an open genome with pSymA, which contains many nodulation- and nitrogen fixation-related genes. This may explain previous observations that horizontal gene transfer has a greater effect on the content of pSymA than pSymB, or the chromosome, and why some rhizobia show unstable nodulation phenotypes on legume hosts.

Loading

Article metrics loading...

/content/journal/mgen/10.1099/mgen.0.000174
2018-04-19
2024-03-28
Loading full text...

Full text loading...

/deliver/fulltext/mgen/4/5/mgen000174.html?itemId=/content/journal/mgen/10.1099/mgen.0.000174&mimeType=html&fmt=ahah

References

  1. Galibert F, Finan TM, Long SR, Puhler A, Abola P et al. The composite genome of the legume symbiont Sinorhizobium meliloti . Science 2001; 293:668–672 http://science.sciencemag.org/content/293/5530/668 (accessed 15 June 2017) [View Article][PubMed]
    [Google Scholar]
  2. Harrison PW, Lower RP, Kim NK, Young JP. Introducing the bacterial 'chromid': not a chromosome, not a plasmid. Trends Microbiol 2010; 18:141–148 [View Article][PubMed]
    [Google Scholar]
  3. Finan TM, Weidner S, Wong K, Buhrmester J, Chain P et al. The complete sequence of the 1,683-kb pSymB megaplasmid from the N2-fixing endosymbiont Sinorhizobium meliloti . Proc Natl Acad Sci USA 2001; 98:9889–9894 [View Article][PubMed]
    [Google Scholar]
  4. Dicenzo GC, MacLean AM, Milunovic B, Golding GB, Finan TM. Examination of prokaryotic multipartite genome evolution through experimental genome reduction. PLoS Genet 2014; 10:e1004742 [View Article][PubMed]
    [Google Scholar]
  5. Galardini M, Pini F, Bazzicalupo M, Biondi EG, Mengoni A. Replicon-dependent bacterial genome evolution: the case of Sinorhizobium meliloti . Genome Biol Evol 2013; 5:542–558 [View Article]
    [Google Scholar]
  6. Dicenzo GC, Checcucci A, Bazzicalupo M, Mengoni A, Viti C et al. Metabolic modelling reveals the specialization of secondary replicons for niche adaptation in Sinorhizobium meliloti . Nat Commun 2016; 7:12219 [View Article][PubMed]
    [Google Scholar]
  7. Kuhn S, Stiens M, Pühler A, Schlüter A. Prevalence of pSmeSM11a-like plasmids in indigenous Sinorhizobium meliloti strains isolated in the course of a field release experiment with genetically modified S. meliloti strains. FEMS Microbiol Ecol 2008; 63:118–131 [View Article]
    [Google Scholar]
  8. Mazur A, Koper P. Rhizobial plasmids — replication, structure and biological role. Open Life Sci 2012; 7:571–586 [View Article]
    [Google Scholar]
  9. Crook MB, Lindsay DP, Biggs MB, Bentley JS, Price JC et al. Rhizobial plasmids that cause impaired symbiotic nitrogen fixation and enhanced host invasion. Mol Plant Microbe Interact 2012; 25:1026–1033 [View Article][PubMed]
    [Google Scholar]
  10. Epstein B, Branca A, Mudge J, Bharti AK, Briskine R et al. Population genomics of the facultatively mutualistic bacteria Sinorhizobium meliloti and S. medicae . PLoS Genet 2012; 8:e1002868 [View Article][PubMed]
    [Google Scholar]
  11. Epstein B, Sadowsky MJ, Tiffin P. Selection on horizontally transferred and duplicated genes in Sinorhizobium (Ensifer), the root-nodule symbionts of medicago. Genome Biol Evol 2014; 6:1199–1209 [View Article][PubMed]
    [Google Scholar]
  12. Herring CD, Palsson . An evaluation of comparative genome sequencing (CGS) by comparing two previously-sequenced bacterial genomes. BMC Genomics 2007; 8:274 [View Article][PubMed]
    [Google Scholar]
  13. Galardini M, Mengoni A, Brilli M, Pini F, Fioravanti A et al. Exploring the symbiotic pangenome of the nitrogen-fixing bacterium Sinorhizobium meliloti . BMC Genomics 2011; 12:235 [View Article]
    [Google Scholar]
  14. Rhoads A, Au KF. PacBio sequencing and its applications. Genomics Proteomics Bioinformatics 2015; 13:278–289 [View Article][PubMed]
    [Google Scholar]
  15. Frost LS, Leplae R, Summers AO, Toussaint A. Mobile genetic elements: the agents of open source evolution. Nat Rev Microbiol 2005; 3:722–732 [View Article]
    [Google Scholar]
  16. Guo X, Flores M, Mavingui P, Fuentes SI, Hernández G et al. Natural genomic design in Sinorhizobium meliloti: novel genomic architectures. Genome Res 2003; 13:1810–1817 [View Article][PubMed]
    [Google Scholar]
  17. Stiens M, Schneiker S, Pühler A, Schlüter A. Sequence analysis of the 181-kb accessory plasmid pSmeSM11b, isolated from a dominant Sinorhizobium meliloti strain identified during a long-term field release experiment. FEMS Microbiol Lett 2007; 271:297–309 [View Article][PubMed]
    [Google Scholar]
  18. Sugawara M, Epstein B, Badgley BD, Unno T, Xu L et al. Comparative genomics of the core and accessory genomes of 48 Sinorhizobium strains comprising five genospecies. Genome Biol 2013; 14:R17 [View Article][PubMed]
    [Google Scholar]
  19. Beringer JE. R factor transfer in Rhizobium leguminosarum . J Gen Microbiol 1974; 84:188–198 [View Article][PubMed]
    [Google Scholar]
  20. Chin CS, Alexander DH, Marks P, Klammer AA, Drake J et al. Nonhybrid, finished microbial genome assemblies from long-read SMRT sequencing data. Nat Methods 2013; 10:563–569 [View Article][PubMed]
    [Google Scholar]
  21. Krumsiek J, Arnold R, Rattei T. Gepard: a rapid and sensitive tool for creating dotplots on genome scale. Bioinformatics 2007; 23:1026–1028 [View Article][PubMed]
    [Google Scholar]
  22. Walker BJ, Abeel T, Shea T, Priest M, Abouelliel A et al. Pilon: an integrated tool for comprehensive microbial variant detection and genome assembly improvement. PLoS One 2014; 9:e112963 [View Article][PubMed]
    [Google Scholar]
  23. Li H, Durbin R. Fast and accurate long-read alignment with Burrows–Wheeler transform. Bioinformatics 2010; 26:589–595 [View Article][PubMed]
    [Google Scholar]
  24. Li H, Handsaker B, Wysoker A, Fennell T, Ruan J et al. The Sequence Alignment/Map format and SAMtools. Bioinformatics 2009; 25:2078–2079 [View Article][PubMed]
    [Google Scholar]
  25. Hyatt D, Chen GL, Locascio PF, Land ML, Larimer FW et al. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics 2010; 11:119 [View Article][PubMed]
    [Google Scholar]
  26. Sallet E, Roux B, Sauviac L, Jardinaud MF, Carrère S et al. Next-generation annotation of prokaryotic genomes with EuGene-P: application to Sinorhizobium meliloti 2011. DNA Res 2013; 20:339–354 [View Article][PubMed]
    [Google Scholar]
  27. Kurtz S, Phillippy A, Delcher AL, Smoot M, Shumway M et al. Versatile and open software for comparing large genomes. Genome Biol 2004; 5:R12 [View Article][PubMed]
    [Google Scholar]
  28. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J et al. BLAST+: architecture and applications. BMC Bioinformatics 2009; 10:421 [View Article][PubMed]
    [Google Scholar]
  29. Gibbons TR, Mount SM, Cooper ED, Delwiche CF. Evaluation of BLAST-based edge-weighting metrics used for homology inference with the Markov Clustering algorithm. BMC Bioinformatics 2015; 16:218 [View Article][PubMed]
    [Google Scholar]
  30. Guhlin J, Silverstein KAT, Zhou P, Tiffin P, Young ND. ODG: Omics database generator – a tool for generating, querying, and analyzing multi-omics comparative databases to facilitate biological understanding. BMC Bioinformatics 2017; 18:367 [View Article][PubMed]
    [Google Scholar]
  31. Vallenet D, Calteau A, Cruveiller S, Gachet M, Lajus A et al. MicroScope in 2017: an expanding and evolving integrated resource for community expertise of microbial genomes. Nucleic Acids Res 2017; 45:D517–D528 [View Article][PubMed]
    [Google Scholar]
  32. Miele V, Penel S, Duret L. Ultra-fast sequence clustering from similarity networks with SiLiX. BMC Bioinformatics 2011; 12:116 [View Article][PubMed]
    [Google Scholar]
  33. Huerta-Cepas J, Szklarczyk D, Forslund K, Cook H, Heller D et al. eggNOG 4.5: a hierarchical orthology framework with improved functional annotations for eukaryotic, prokaryotic and viral sequences. Nucleic Acids Res 2016; 44:D286–D293 [View Article][PubMed]
    [Google Scholar]
  34. Finn RD, Clements J, Eddy SR. HMMER web server: interactive sequence similarity searching. Nucleic Acids Res 2011; 39:W29–W37 [View Article][PubMed]
    [Google Scholar]
  35. Enright AJ, van Dongen S, Ouzounis CA. An efficient algorithm for large-scale detection of protein families. Nucleic Acids Res 2002; 30:1575–1584 [View Article][PubMed]
    [Google Scholar]
  36. Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol 2013; 30:772–780 [View Article][PubMed]
    [Google Scholar]
  37. Capella-Gutiérrez S, Silla-Martínez JM, Gabaldón T. trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 2009; 25:1972–1973 [View Article][PubMed]
    [Google Scholar]
  38. Principal component analysis. Chemom Intell Lab Syst 1987; 2:37–52
    [Google Scholar]
  39. Paradis E, Claude J, Strimmer K. APE: analyses of phylogenetics and evolution in R language. Bioinformatics 2004; 20:289–290 [View Article]
    [Google Scholar]
  40. Tamura K, Nei M. Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Mol Biol Evol 1993; 10:512–526 [View Article][PubMed]
    [Google Scholar]
  41. Mantel N. The detection of disease clustering and a generalized regression approach. Cancer Res 1967; 27:209–220[PubMed]
    [Google Scholar]
  42. Dereeper A, Audic S, Claverie JM, Blanc G. BLAST-EXPLORER helps you building datasets for phylogenetic analysis. BMC Evol Biol 2010; 10:8 [View Article][PubMed]
    [Google Scholar]
  43. Dereeper A, Guignon V, Blanc G, Audic S, Buffet S et al. Phylogeny.fr: robust phylogenetic analysis for the non-specialist. Nucleic Acids Res 2008; 36:W465–W469 [View Article][PubMed]
    [Google Scholar]
  44. Edgar RC. MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 2004; 32:1792–1797 [View Article][PubMed]
    [Google Scholar]
  45. Castresana J. Selection of conserved blocks from multiple alignments for their use in phylogenetic analysis. Mol Biol Evol 2000; 17:540–552 [View Article][PubMed]
    [Google Scholar]
  46. Guindon S, Gascuel O, A Simple RB. A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Syst Biol 2003; 52:696–704 [View Article][PubMed]
    [Google Scholar]
  47. Anisimova M, Gascuel O, Sullivan J. Approximate likelihood-ratio test for branches: A fast, accurate, and powerful alternative. Syst Biol 2006; 55:539–552 [View Article][PubMed]
    [Google Scholar]
  48. Chevenet F, Brun C, Bañuls AL, Jacq B, Christen R. TreeDyn: towards dynamic graphics and annotations for analyses of trees. BMC Bioinformatics 2006; 7:439 [View Article][PubMed]
    [Google Scholar]
  49. Martínez-Abarca F, Martínez-Rodríguez L, López-Contreras JA, Jiménez-Zurdo JI, Toro N. Complete genome sequence of the alfalfa symbiont Sinorhizobium/Ensifer meliloti Strain GR4. Genome Announc 2013; 1:e00174-12-e00174-12e00174-12 [View Article][PubMed]
    [Google Scholar]
  50. Schneiker-Bekel S, Wibberg D, Bekel T, Blom J, Linke B et al. The complete genome sequence of the dominant Sinorhizobium meliloti field isolate SM11 extends the S. meliloti pan-genome. J Biotechnol 2011; 155:20–33 [View Article][PubMed]
    [Google Scholar]
  51. Cevallos MA, Cervantes-Rivera R, Gutiérrez-Ríos RM. The repABC plasmid family. Plasmid 2008; 60:19–37 [View Article][PubMed]
    [Google Scholar]
  52. van DSM. Graph clustering by flow simulation; 2000 https://dspace.library.uu.nl/handle/1874/848 (accessed 13 November 2017)
  53. Manen D, Upegui-Gonzalez LC, Caro L. Monomers and dimers of the RepA protein in plasmid pSC101 replication: domains in RepA. Proc Natl Acad Sci USA 1992; 89:8923–8927 [View Article][PubMed]
    [Google Scholar]
  54. Hiei Y, Ohta S, Komari T, Kumashiro T. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J 1994; 6:271–282 [View Article][PubMed]
    [Google Scholar]
  55. Ellis JG, Murphy PJ. Four new opines from crown gall tumours – Their detection and properties. MGG Mol Gen Genet 1981; 181:36–43 [View Article]
    [Google Scholar]
  56. Moore LW, Chilton WS, Canfield ML. Diversity of opines and opine-catabolizing bacteria isolated from naturally occurring crown gall tumors. Appl Environ Microbiol 1997; 63:201–207[PubMed]
    [Google Scholar]
  57. Mavingui P, Flores M, Guo X, Dávila G, Perret X et al. Dynamics of genome architecture in Rhizobium sp. strain NGR234. J Bacteriol 2002; 184:171–176 [View Article][PubMed]
    [Google Scholar]
  58. Dicenzo G, Milunovic B, Cheng J, Finan TM. The tRNAarg gene and engA are essential genes on the 1.7-Mb pSymB megaplasmid of Sinorhizobium meliloti and were translocated together from the chromosome in an ancestral strain. J Bacteriol 2013; 195:202–212 [View Article][PubMed]
    [Google Scholar]
  59. Wei M, Zhang JJ, Liu H, Wang SJ, Fu H et al. A transposable class I composite transposon carrying mph (methyl parathion hydrolase) from Pseudomonas sp. strain WBC-3. FEMS Microbiol Lett 2009; 292:85–91 [View Article][PubMed]
    [Google Scholar]
  60. Mijnendonckx K, Provoost A, Monsieurs P, Leys N, Mergeay M et al. Insertion sequence elements in Cupriavidus metallidurans CH34: distribution and role in adaptation. Plasmid 2011; 65:193–203 [View Article][PubMed]
    [Google Scholar]
  61. Thomas CM, Nielsen KM. Mechanisms of, and barriers to, horizontal gene transfer between bacteria. Nat Rev Microbiol 2005; 3:711–721 [View Article][PubMed]
    [Google Scholar]
  62. Darmon E, Leach DRF. Bacterial genome instability. Microbiol Mol Biol Rev 2014; 78:1–39 [View Article]
    [Google Scholar]
  63. Treangen TJ, Abraham AL, Touchon M, Rocha EP. Genesis, effects and fates of repeats in prokaryotic genomes. FEMS Microbiol Rev 2009; 33:539–571 [View Article][PubMed]
    [Google Scholar]
  64. Dillon MM, Sung W, Lynch M, Cooper VS. The rate and molecular spectrum of spontaneous mutations in the GC-Rich multichromosome genome of Burkholderia cenocepacia . Genetics 2015; 200:935–946 [View Article][PubMed]
    [Google Scholar]
  65. Daubin V, Szöllősi GJ. Horizontal gene transfer and the history of life. Cold Spring Harb Perspect Biol 2016; 8:a018036 [View Article][PubMed]
    [Google Scholar]
  66. Romero D, Martínez-Salazar J, Girard L, Brom S, Dávilla G et al. Discrete amplifiable regions (amplicons) in the symbiotic plasmid of Rhizobium etli CFN42. J Bacteriol 1995; 177:973–980 [View Article][PubMed]
    [Google Scholar]
  67. Romero D, Brom S, Martínez-Salazar J, Girard ML, Palacios R et al. Amplification and deletion of a nod-nif region in the symbiotic plasmid of Rhizobium phaseoli . J Bacteriol 1991; 173:2435–2441 [View Article][PubMed]
    [Google Scholar]
  68. Barnett MJ, Kahn ML. Psyma of Sinorhizobium meliloti: nitrogen fixation and more. Genomes and Genomics of Nitrogen-fixing Organisms Berlin/Heidelberg: Springer-Verlag; pp. 113–132
    [Google Scholar]
  69. Nelson MS, Chun CL, Sadowsky MJ. Type IV effector proteins involved in the MedicagoSinorhizobium symbiosis. Mol Plant-Microbe Interact 2017; 30:28–34 [View Article]
    [Google Scholar]
  70. Hubber A, Vergunst AC, Sullivan JT, Hooykaas PJ, Ronson CW. Symbiotic phenotypes and translocated effector proteins of the Mesorhizobium loti strain R7A VirB/D4 type IV secretion system. Mol Microbiol 2004; 54:561–574 [View Article][PubMed]
    [Google Scholar]
  71. Farrand SK, van Berkum PB, Oger P. Agrobacterium is a definable genus of the family Rhizobiaceae . Int J Syst Evol Microbiol 2003; 53:1681–1687 [View Article][PubMed]
    [Google Scholar]
  72. van Veen RJM, den Dulk-Ras H, Schilperoort RA, Hooykaas PJJ. Ti plasmid containing Rhizobium meliloti are non-tumorigenic on plants, despite proper virulence gene induction and T-strand formation. Arch Microbiol 1989; 153:85–89 [View Article]
    [Google Scholar]
  73. Hooykaas PJJ, Klapwijk PM, Nuti MP, Schilperoort RA, Rorsch A. Transfer of the Agrobacterium tumefaciens TI plasmid to avirulent agrobacteria and to Rhizobium ex planta . J Gen Microbiol 1977; 98:477–484 [View Article]
    [Google Scholar]
  74. Paulus F, Otten L. Functional and mutated agrocinopine synthase genes on octopine T-DNAs. Mol Plant Microbe Interact 1993; 6:393–402 [View Article][PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/mgen/10.1099/mgen.0.000174
Loading
/content/journal/mgen/10.1099/mgen.0.000174
Loading

Data & Media loading...

Supplements

Supplementary File 3

PDF

Supplementary File 2

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error