1887

Abstract

Purpose. We resequenced the genome of Clostridium difficile 630Δerm (DSM 28645), a model strain commonly used for the generation of insertion mutants.

Methodology. The genome sequence was obtained by a combination of single-molecule real-timeand Illumina sequencing technology.

Results. Detailed manual curation and comparison to the previously published genomic sequence revealed sequence differences including inverted regions and the presence of plasmid pCD630. Manual curation of our previously deposited genome sequence of the parental strain 630 (DSM 27543) led to an improved genome sequence. In addition, the sequence of the transposon Tn5397 was completely identified. We manually revised the current manual annotation of the initial sequence of strain 630 and modified either gene names, gene product names or assigned EC numbers of 57 % of genes. The number of hypothetical and conserved hypothetical proteins was reduced by 152. This annotation was used as a template to annotate the most recent genome sequences of the strains 630Δerm and 630.

Conclusion. Based on the genomic analysis, several new metabolic features of C. difficile are proposed and could be supported by literature and subsequent experiments.

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2017-03-23
2019-10-18
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References

  1. Wüst J, Sullivan NM, Hardegger U, Wilkins TD. Investigation of an outbreak of antibiotic-associated colitis by various typing methods. J Clin Microbiol 1982;16:1096–1101[PubMed]
    [Google Scholar]
  2. Hussain HA, Roberts AP, Mullany P. Generation of an erythromycin-sensitive derivative of Clostridium difficile strain 630 (630Δerm) and demonstration that the conjugative transposon Tn916ΔE enters the genome of this strain at multiple sites. J Med Microbiol 2005;54:137–141 [CrossRef][PubMed]
    [Google Scholar]
  3. Heap JT, Pennington OJ, Cartman ST, Carter GP, Minton NP. The ClosTron: a universal gene knock-out system for the genus Clostridium. J Microbiol Methods 2007;70:452–464 [CrossRef][PubMed]
    [Google Scholar]
  4. Sebaihia M, Wren BW, Mullany P, Fairweather NF, Minton N et al. The multidrug-resistant human pathogen Clostridium difficile has a highly mobile, mosaic genome. Nat Genet 2006;38:779–786 [CrossRef][PubMed]
    [Google Scholar]
  5. Monot M, Boursaux-Eude C, Thibonnier M, Vallenet D, Moszer I et al. Reannotation of the genome sequence of Clostridium difficile strain 630. J Med Microbiol 2011;60:1193–1199 [CrossRef][PubMed]
    [Google Scholar]
  6. Pettit LJ, Browne HP, Yu L, Smits WK, Fagan RP et al. Functional genomics reveals that Clostridium difficile Spo0A coordinates sporulation, virulence and metabolism. BMC Genomics 2014;15:160 [CrossRef][PubMed]
    [Google Scholar]
  7. Riedel T, Bunk B, Thürmer A, Spröer C, Brzuszkiewicz E et al. Genome resequencing of the virulent and multidrug-resistant reference strain Clostridium difficile 630. Genome Announc 2015;3:e00276-15 [CrossRef][PubMed]
    [Google Scholar]
  8. Van Eijk E, Anvar SY, Browne HP, Leung WY, Frank J et al. Complete genome sequence of the Clostridium difficile laboratory strain 630Δerm reveals differences from strain 630, including translocation of the mobile element CTn5. BMC Genomics 2015;16:31 [CrossRef][PubMed]
    [Google Scholar]
  9. Collery MM, Kuehne SA, Mcbride SM, Kelly ML, Monot M et al. What's a SNP between friends: the influence of single nucleotide polymorphisms on virulence and phenotypes of Clostridium difficile strain 630 and derivatives. Virulence 2016;1–15 [CrossRef][PubMed]
    [Google Scholar]
  10. Riedel T, Bunk B, Wittmann J, Thürmer A, Spröer C et al. Complete genome sequence of the Clostridium difficile type strain DSM 1296T. Genome Announc 2015;3:e01186-15 [CrossRef][PubMed]
    [Google Scholar]
  11. Neumann-Schaal M, Hofmann JD, Will SE, Schomburg D. Time-resolved amino acid uptake of Clostridium difficile 630Δerm and concomitant fermentation product and toxin formation. BMC Microbiol 2015;15:281 [CrossRef][PubMed]
    [Google Scholar]
  12. Li H, Durbin R. Fast and accurate long-read alignment with Burrows-Wheeler transform. Bioinformatics 2010;26:589–595 [CrossRef][PubMed]
    [Google Scholar]
  13. Robinson JT, Thorvaldsdóttir H, Winckler W, Guttman M, Lander ES et al. Integrative Genomics Viewer. Nat Biotechnol 2011;29:24–26 [CrossRef][PubMed]
    [Google Scholar]
  14. Thorvaldsdóttir H, Robinson JT, Mesirov JP. Integrative Genomics Viewer (IGV): high-performance genomics data visualization and exploration. Brief Bioinform 2013;14:178–192 [CrossRef][PubMed]
    [Google Scholar]
  15. Darling ACE, Mau B, Blattner FR, Perna NT. Mauve: multiple alignment of conserved genomic sequence with rearrangements. Genome Res 2004;14:1394–1403 [CrossRef][PubMed]
    [Google Scholar]
  16. Darling AE, Treangen TJ, Messeguer X, Perna NT. Analyzing patterns of microbial evolution using the mauve genome alignment system. Methods Mol Biol Clifton NJ 2007;396:135–152[CrossRef]
    [Google Scholar]
  17. Darling AE, Mau B, Perna NT. progressive mauve: multiple genome alignment with gene gain, loss and rearrangement. PLoS One 2010;5:e11147 [CrossRef][PubMed]
    [Google Scholar]
  18. Mullany P, Pallen M, Wilks M, Stephen JR, Tabaqchali S. A group II intron in a conjugative transposon from the gram-positive bacterium, Clostridium difficile. Gene 1996;174:145–150 [CrossRef][PubMed]
    [Google Scholar]
  19. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J et al. BLAST+: architecture and applications. BMC Bioinformatics 2009;10:421 [CrossRef][PubMed]
    [Google Scholar]
  20. Bannert C, Welfle A, Aus dem Spring C, Schomburg D. BrEPS: a flexible and automatic protocol to compute enzyme-specific sequence profiles for functional annotation. BMC Bioinformatics 2010;11:589 [CrossRef][PubMed]
    [Google Scholar]
  21. Jones P, Binns D, Chang H-Y, Fraser M, Li W et al. InterProScan 5: genome-scale protein function classification. Bioinformatics 2014;30:1236–1240 [CrossRef][PubMed]
    [Google Scholar]
  22. Seemann T. Prokka: rapid prokaryotic genome annotation. Bioinformatics 2014;30:2068–2069 [CrossRef]
    [Google Scholar]
  23. Zech H, Thole S, Schreiber K, Kalhöfer D, Voget S et al. Growth phase-dependent global protein and metabolite profiles of Phaeobacter gallaeciensis strain DSM 17395, a member of the marine Roseobacter-clade. Proteomics 2009;9:3677–3697 [CrossRef][PubMed]
    [Google Scholar]
  24. Kruskal WH, Wallis WA. Use of ranks in one-criterion variance analysis. J Am Stat Assoc 1952;47:583–621 [CrossRef]
    [Google Scholar]
  25. Benjamini Y, Yekutieli D. The control of the false discovery rate in multiple testing under dependency. Ann Stat 2001;29:1165–1188[CrossRef]
    [Google Scholar]
  26. Wolf J, Stark H, Fafenrot K, Albersmeier A, Pham TK et al. A systems biology approach reveals major metabolic changes in the thermoacidophilic archaeon Sulfolobus solfataricus in response to the carbon source L-fucose versus D-glucose. Mol Microbiol 2016;102:882–908 [CrossRef][PubMed]
    [Google Scholar]
  27. Emerson JE, Reynolds CB, Fagan RP, Shaw HA, Goulding D et al. A novel genetic switch controls phase variable expression of CwpV, a Clostridium difficile cell wall protein. Mol Microbiol 2009;74:541–556 [CrossRef][PubMed]
    [Google Scholar]
  28. Stabler RA, Valiente E, Dawson LF, He M, Parkhill J et al. In-depth genetic analysis of Clostridium difficile PCR-ribotype 027 strains reveals high genome fluidity including point mutations and inversions. Gut Microbes 2010;1:269–276 [CrossRef][PubMed]
    [Google Scholar]
  29. Povelainen M, Miasnikov AN. Production of xylitol by metabolically engineered strains of Bacillus subtilis. J Biotechnol 2007;128:24–31 [CrossRef][PubMed]
    [Google Scholar]
  30. Faulds-Pain A, Twine SM, Vinogradov E, Strong PCR, Dell A et al. The post-translational modification of the Clostridium difficile flagellin affects motility, cell surface properties and virulence. Mol Microbiol 2014;94:272–289 [CrossRef][PubMed]
    [Google Scholar]
  31. Li F, Hagemeier CH, Seedorf H, Gottschalk G, Thauer RK. Re-citrate synthase from Clostridium kluyveri is phylogenetically related to homocitrate synthase and isopropylmalate synthase rather than to Si-citrate synthase. J Bacteriol 2007;189:4299–4304 [CrossRef][PubMed]
    [Google Scholar]
  32. Wang N-C, Lee C-Y. Molecular cloning of the aspartate 4-decarboxylase gene from Pseudomonas sp. ATCC 19121 and characterization of the bifunctional recombinant enzyme. Appl Microbiol Biotechnol 2006;73:339–348 [CrossRef][PubMed]
    [Google Scholar]
  33. Coleman JP, Hudson LL, Adams MJ. Characterization and regulation of the NADP-linked 7α-hydroxysteroid dehydrogenase gene from Clostridium sordellii. J Bacteriol 1994;176:4865–4874 [CrossRef][PubMed]
    [Google Scholar]
  34. Midtvedt T, Norman A. Bile acid transformations by microbial strains belonging to genera found in intestinal contents. Acta Pathol Microbiol Scand 1967;71:629–638 [CrossRef][PubMed]
    [Google Scholar]
  35. Sorg JA, Sonenshein AL. Bile salts and glycine as cogerminants for clostridium difficile spores. J Bacteriol 2008;190:2505–2512 [CrossRef][PubMed]
    [Google Scholar]
  36. Wilson KH. Efficiency of various bile salt preparations for stimulation of Clostridium difficile spore germination. J Clin Microbiol 1983;18:1017–1019[PubMed]
    [Google Scholar]
  37. Ridlon JM, Kang D-J, Hylemon PB. Bile salt biotransformations by human intestinal bacteria. J Lipid Res 2006;47:241–259 [CrossRef][PubMed]
    [Google Scholar]
  38. Buffie CG, Bucci V, Stein RR, Mckenney PT, Ling L et al. Precision microbiome reconstitution restores bile acid mediated resistance to Clostridium difficile. Nature 2015;517:205–208 [CrossRef][PubMed]
    [Google Scholar]
  39. Sorg JA. Microbial bile acid metabolic clusters: the bouncers at the bar. Cell Host Microbe 2014;16:551–552 [CrossRef][PubMed]
    [Google Scholar]
  40. Kim J, Darley D, Selmer T, Buckel W. Characterization of (R)-2-Hydroxyisocaproate dehydrogenase and a family III coenzyme A transferase involved in reduction of L-leucine to isocaproate by Clostridium difficile. Appl Environ Microbiol 2006;72:6062–6069 [CrossRef][PubMed]
    [Google Scholar]
  41. Weghoff MC, Bertsch J, Müller V. A novel mode of lactate metabolism in strictly anaerobic bacteria. Environ Microbiol 2015;17:670–677 [CrossRef][PubMed]
    [Google Scholar]
  42. Desguin B, Goffin P, Viaene E, Kleerebezem M, Martin-Diaconescu V et al. Lactate racemase is a nickel-dependent enzyme activated by a widespread maturation system. Nat Commun 2014;5:3615 [CrossRef][PubMed]
    [Google Scholar]
  43. Karasawa T, Ikoma S, Yamakawa K, Nakamura S. A defined growth medium for Clostridium difficile. Microbiology 1995;141:371–375 [CrossRef]
    [Google Scholar]
  44. Van den Heuvel RHH, Ferrari D, Bossi RT, Ravasio S, Curti B et al. Structural studies on the synchronization of catalytic centers in glutamate synthase. J Biol Chem 2002;277:24579–24583 [CrossRef][PubMed]
    [Google Scholar]
  45. Girinathan BP, Braun SE, Govind R. Clostridium difficile glutamate dehydrogenase is a secreted enzyme that confers resistance to H2O2. Microbiology 2014;160:47–55 [CrossRef][PubMed]
    [Google Scholar]
  46. O'Brien JR, Raynaud C, Croux C, Girbal L, Soucaille P et al. Insight into the mechanism of the B12 - independent glycerol dehydratase from Clostridium butyricum: preliminary biochemical and structural characterization. Biochemistry 2004;43:4635–4645 [CrossRef][PubMed]
    [Google Scholar]
  47. Raynaud C, Sarçabal P, Meynial-Salles I, Croux C, Soucaille P. Molecular characterization of the 1,3-propanediol (1,3-PD) operon of Clostridium butyricum. Proc Natl Acad Sci USA 2003;100:5010–5015 [CrossRef][PubMed]
    [Google Scholar]
  48. Stickland LH. Studies in the metabolism of the strict anaerobes (genus Clostridium). Biochem J 1934;28:1746–1759 [CrossRef]
    [Google Scholar]
  49. Vogel HJ, Davis BD. Glutamic γ-semialdehyde and Δ1-pyrroline-5-carboxylic acid, intermediates in the biosynthesis of proline. J Am Chem Soc 1952;74:109–112 [CrossRef]
    [Google Scholar]
  50. Williams I, Frank L. Improved chemical synthesis and enzymatic assay of Δ1-pyrroline-5-carboxylic acid. Anal Biochem 1975;64:85–97 [CrossRef][PubMed]
    [Google Scholar]
  51. Jackson S, Calos M, Myers A, Self WT. Analysis of proline reduction in the nosocomial pathogen Clostridium difficile. J Bacteriol 2006;188:8487–8495 [CrossRef][PubMed]
    [Google Scholar]
  52. Mai X, Adams MW. Indolepyruvate ferredoxin oxidoreductase from the hyperthermophilic archaeon Pyrococcus furiosus. A new enzyme involved in peptide fermentation. J Biol Chem 1994;269:16726–16732[PubMed]
    [Google Scholar]
  53. Tersteegen A, Linder D, Thauer RK, Hedderich R. Structures and functions of four anabolic 2-oxoacid oxidoreductases in Methanobacterium thermoautotrophicum. Eur J Biochem 1997;244:862–868 [CrossRef]
    [Google Scholar]
  54. Pereira FL, Oliveira Júnior CA, Silva ROS, Dorella FA, Carvalho AF et al. Complete genome sequence of Peptoclostridium difficile strain Z31. Gut Pathog 2016;8:11 [CrossRef]
    [Google Scholar]
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