1887

Abstract

(formerly ) colonizes the gastrointestinal tract following disruption of the microbiota and can initiate a spectrum of clinical manifestations ranging from asymptomatic to life-threatening colitis. Following antibiotic treatment, luminal oxygen concentrations increase, exposing gut microbes to potentially toxic reactive oxygen species. Though typically regarded as a strict anaerobe, can grow at low oxygen concentrations. How this bacterium adapts to a microaerobic environment and whether those responses to oxygen are conserved amongst strains is not entirely understood. Here, two strains (630 and CD196) were cultured in 1.5% oxygen and the transcriptional response to long-term oxygen exposure was evaluated via RNA-sequencing. During growth in a microaerobic environment, several genes predicted to protect against oxidative stress were upregulated, including those for rubrerythrins and rubredoxins. Transcription of genes involved in metal homeostasis was also positively correlated with increased oxygen levels and these genes were amongst the most differentially transcribed. To directly compare the transcriptional landscape between strains, a ‘consensus-genome’ was generated. On the basis of the identified conserved genes, basal transcriptional differences as well as variations in the response to oxygen were evaluated. While several responses were similar between the strains, there were significant differences in the abundance of transcripts involved in amino acid and carbohydrate metabolism. Furthermore, intracellular metal concentrations significantly varied both in an oxygen-dependent and oxygen-independent manner. Overall, these results indicate that adapts to grow in a low oxygen environment through transcriptional changes, though the specific strategy employed varies between strains.

Funding
This study was supported by the:
  • California State University, Sacramento
    • Principle Award Recipient: JhoanaRodriguez
  • National Institute of Allergy and Infectious Diseases (Award F32AI157215)
    • Principle Award Recipient: AndyWeiss
  • National Institute of Environmental Health Sciences (Award T32ES007028)
    • Principle Award Recipient: AndyWeiss
  • American Heart Association (Award 18POST33990262)
    • Principle Award Recipient: AndyWeiss
  • American Heart Association (Award 18POST34030426)
    • Principle Award Recipient: WilliamN. Beavers
  • Jane Coffin Childs Memorial Fund for Medical Research
    • Principle Award Recipient: ChristopherA. Lopez
  • Burroughs Wellcome Fund
    • Principle Award Recipient: ChristopherA. Lopez
  • National Institute of Diabetes and Digestive and Kidney Diseases (Award DK058404)
    • Principle Award Recipient: EricP Skaar
  • National Institute of Allergy and Infectious Diseases (Award R01 AI118089)
    • Principle Award Recipient: EricP Skaar
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License.
Loading

Article metrics loading...

/content/journal/mgen/10.1099/mgen.0.000738
2021-12-15
2022-01-29
Loading full text...

Full text loading...

/deliver/fulltext/mgen/7/12/mgen000738.html?itemId=/content/journal/mgen/10.1099/mgen.0.000738&mimeType=html&fmt=ahah

References

  1. Rodriguez Diaz C, Seyboldt C, Rupnik M, Non-human C. Non-human C. difficile reservoirs and sources: animals, food, environment. Adv Exp Med Biol 2018; 1050:227–243 [View Article] [PubMed]
    [Google Scholar]
  2. Knight DR, Elliott B, Chang BJ, Perkins TT, Riley TV. Diversity and evolution in the genome of Clostridium difficile. Clin Microbiol Rev 2015; 28:721–741 [View Article] [PubMed]
    [Google Scholar]
  3. Abt MC, McKenney PT, Pamer EG. Clostridium difficile colitis: pathogenesis and host defence. Nat Rev Microbiol 2016; 14:609–620 [View Article] [PubMed]
    [Google Scholar]
  4. Chandrasekaran R, Lacy DB. The role of toxins in Clostridium difficile infection. FEMS Microbiol Rev 2017; 41:723–750 [View Article] [PubMed]
    [Google Scholar]
  5. Czepiel J, Dróżdż M, Pituch H, Kuijper EJ, Perucki W et al. Clostridium difficile infection: review. Eur J Clin Microbiol Infect Dis 2019; 38:1211–1221 [View Article] [PubMed]
    [Google Scholar]
  6. Eyre DW, Griffiths D, Vaughan A, Golubchik T, Acharya M et al. Asymptomatic Clostridium difficile colonisation and onward transmission. PLoS One 2013; 8:e78445 [View Article]
    [Google Scholar]
  7. Crobach MJT, Vernon JJ, Loo VG, Kong LY, Péchiné S et al. Understanding Clostridium difficile colonization. Clin Microbiol Rev 2018; 31:e00021-17. [View Article] [PubMed]
    [Google Scholar]
  8. Litvak Y, Byndloss MX, Bäumler AJ. Colonocyte metabolism shapes the gut microbiota. Science 2018; 362:eaat9076 [View Article] [PubMed]
    [Google Scholar]
  9. Donohoe DR, Garge N, Zhang X, Sun W, O’Connell TM et al. The microbiome and butyrate regulate energy metabolism and autophagy in the mammalian colon. Cell Metab 2011; 13:517–526 [View Article] [PubMed]
    [Google Scholar]
  10. Duszka K, Oresic M, Le May C, König J, Wahli W. PPARγ modulates long chain fatty acid processing in the intestinal epithelium. IJMS 2017; 18:2559 [View Article]
    [Google Scholar]
  11. Zheng L, Kelly CJ, Colgan SP. Physiologic hypoxia and oxygen homeostasis in the healthy intestine. A review in the theme: cellular responses to hypoxia. Am J Physiol Cell Physiol 2015; 309:C350–60 [View Article] [PubMed]
    [Google Scholar]
  12. Rivera-Chávez F, Zhang LF, Faber F, Lopez CA, Byndloss MX et al. Depletion of butyrate-producing Clostridia from the gut microbiota drives an aerobic luminal expansion of Salmonella. Cell Host Microbe 2016; 19:443–454 [View Article] [PubMed]
    [Google Scholar]
  13. Theriot CM, Koenigsknecht MJ, Carlson PE Jr, Hatton GE, Nelson AM et al. Antibiotic-induced shifts in the mouse gut microbiome and metabolome increase susceptibility to Clostridium difficile infection. Nat Commun 2014; 5:3114. [View Article] [PubMed]
    [Google Scholar]
  14. Giordano N, Hastie JL, Smith AD, Foss ED, Gutierrez-Munoz DF et al. Cysteine desulfurase IscS2 plays a role in oxygen resistance in Clostridium difficile. Infect Immun 2018; 86:e00326-18. [View Article] [PubMed]
    [Google Scholar]
  15. Fachi JL, Felipe J de S, Pral LP, da Silva BK, Corrêa RO et al. Butyrate protects mice from Clostridium difficile-induced colitis through an HIF-1-dependent mechanism. Cell Reports 2019; 27:750–761 [View Article]
    [Google Scholar]
  16. Emerson JE, Stabler RA, Wren BW, Fairweather NF. Microarray analysis of the transcriptional responses of Clostridium difficile to environmental and antibiotic stress. J Med Microbiol 2008; 57:757–764 [View Article] [PubMed]
    [Google Scholar]
  17. Neumann-Schaal M, Metzendorf NG, Troitzsch D, Nuss AM, Hofmann JD et al. Tracking gene expression and oxidative damage of O2-stressed Clostridioides difficile by a multi-omics approach. Anaerobe 2018; 53:94–107 [View Article] [PubMed]
    [Google Scholar]
  18. Folgosa F, Martins MC, Teixeira M. The multidomain flavodiiron protein from Clostridium difficile 630 is an NADH:oxygen oxidoreductase. Sci Rep 2018; 8:10164. [View Article] [PubMed]
    [Google Scholar]
  19. Giordano N, Hastie JL, Carlson PE. Transcriptomic profiling of Clostridium difficile grown under microaerophillic conditions. Pathog Dis 2018; 76: [View Article] [PubMed]
    [Google Scholar]
  20. 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 [View Article] [PubMed]
    [Google Scholar]
  21. Edwards AN, Suárez JM, McBride SM. Culturing and maintaining Clostridium difficile in an anaerobic environment. JoVE 201379 [View Article]
    [Google Scholar]
  22. Winston JA, Thanissery R, Montgomery SA, Theriot CM. Cefoperazone-treated mouse model of clinically-relevant Clostridium difficile strain R20291. JoVE 2016118 [View Article]
    [Google Scholar]
  23. Lopez CA, Beavers WN, Weiss A, Knippel RJ, Zackular JP et al. The immune protein calprotectin impacts Clostridioides difficile metabolism through zinc limitation. mBio 2019; 10:e02289-19. [View Article] [PubMed]
    [Google Scholar]
  24. Gross MW, Karbach U, Groebe K, Franko AJ, Mueller-Klieser W. Calibration of misonidazole labeling by simultaneous measurement of oxygen tension and labeling density in multicellular spheroids. Int J Cancer 1995; 61:567–573 [View Article] [PubMed]
    [Google Scholar]
  25. Stabler RA, He M, Dawson L, Martin M, Valiente E et al. Comparative genome and phenotypic analysis of Clostridium difficile 027 strains provides insight into the evolution of a hypervirulent bacterium. Genome Biol 2009; 10:R102. [View Article] [PubMed]
    [Google Scholar]
  26. Anjuwon-Foster BR, Maldonado-Vazquez N, Tamayo R, Zhulin IB. Characterization of flagellum and toxin phase variation in Clostridioides difficile ribotype 012 isolates. J Bacteriol 2018; 200:14 [View Article]
    [Google Scholar]
  27. Trzilova D, Anjuwon-Foster BR, Torres Rivera D, Tamayo R. Rho factor mediates flagellum and toxin phase variation and impacts virulence in Clostridioides difficile. PLoS Pathog 2020; 16:e1008708 [View Article] [PubMed]
    [Google Scholar]
  28. Steinberg HD, Snitkin ES. Homologous recombination in Clostridioides difficile mediates diversification of cell surface features and transport systems. mSphere 2020; 5:e00799-20. [View Article] [PubMed]
    [Google Scholar]
  29. Knight DR, Imwattana K, Kullin B, Guerrero-Araya E, Paredes-Sabja D et al. Major genetic discontinuity and novel toxigenic species in Clostridioides difficile taxonomy. elife 2021; 10:e64325. [View Article] [PubMed]
    [Google Scholar]
  30. Kumar N, Browne HP, Viciani E, Forster SC, Clare S et al. Adaptation of host transmission cycle during Clostridium difficile speciation. Nat Genet 2019; 51:1315–1320 [View Article] [PubMed]
    [Google Scholar]
  31. Lopez CA, Miller BM, Rivera-Chávez F, Velazquez EM, Byndloss MX et al. Virulence factors enhance Citrobacter rodentium expansion through aerobic respiration. Science 2016; 353:1249–1253 [View Article] [PubMed]
    [Google Scholar]
  32. Morvan C, Folgosa F, Kint N, Teixeira M, Martin-Verstraete I. Responses of Clostridia to oxygen: from detoxification to adaptive strategies. Environ Microbiol 2021; 23:4112–4125 [View Article] [PubMed]
    [Google Scholar]
  33. Kint N, Alves Feliciano C, Martins MC, Morvan C, Fernandes SF et al. Erratum for Kint et al., “How the anaerobic enteropathogen Clostridioides difficile tolerates low O2 tensions”. mBio 2020; 11:e02678-20. [View Article] [PubMed]
    [Google Scholar]
  34. Gaupp R, Ledala N, Somerville GA. Staphylococcal response to oxidative stress. Front Cell Infect Microbiol 2012; 2:33. [View Article] [PubMed]
    [Google Scholar]
  35. Chiang SM, Schellhorn HE. Regulators of oxidative stress response genes in Escherichia coli and their functional conservation in bacteria. Arch Biochem Biophys 2012; 525:161–169 [View Article] [PubMed]
    [Google Scholar]
  36. Coulter ED. A role for rubredoxin in oxidative stress protection in Desulfovibrio vulgaris: catalytic electron transfer to rubrerythrin and two-iron superoxide reductase. Arch Biochem Biophys 2001; 394:76–86 [View Article] [PubMed]
    [Google Scholar]
  37. Troitzsch D, Zhang H, Dittmann S, Düsterhöft D, Möller TA et al. A point mutation in the transcriptional repressor PerR results in a constitutive oxidative stress response in Clostridioides difficile 630Δerm. mSphere 2021; 6:e00091-21. [View Article] [PubMed]
    [Google Scholar]
  38. Hillmann F, Fischer RJ, Saint-Prix F, Girbal L, Bahl H. PerR acts as a switch for oxygen tolerance in the strict anaerobe Clostridium acetobutylicum. Mol Microbiol 2008; 68:848–860 [View Article] [PubMed]
    [Google Scholar]
  39. Saujet L, Pereira FC, Serrano M, Soutourina O, Monot M et al. Genome-wide analysis of cell type-specific gene transcription during spore formation in Clostridium difficile. PLoS Genet 2013; 9:10e1003756 [View Article] [PubMed]
    [Google Scholar]
  40. Boekhoud IM, Michel AM, Corver J, Jahn D, Smits WK. Redefining the Clostridioides difficile σB Regulon: σB activates genes involved in detoxifying radicals that can result from the exposure to antimicrobials and hydrogen peroxide. mSphere 2020; 5:e00728-20. [View Article] [PubMed]
    [Google Scholar]
  41. May A, Hillmann F, Riebe O, Fischer RJ, Bahl H. A rubrerythrin-like oxidative stress protein of Clostridium acetobutylicum is encoded by a duplicated gene and identical to the heat shock protein Hsp21. FEMS Microbiol Lett 2004; 238:249–254 [View Article] [PubMed]
    [Google Scholar]
  42. Knippel RJ, Wexler AG, Miller JM, Beavers WN, Weiss A et al. Clostridioides difficile senses and hijacks host heme for incorporation into an oxidative stress defense system. Cell Host & Microbe 2020; 28:411–421 [View Article]
    [Google Scholar]
  43. Lu J, Holmgren A. The thioredoxin antioxidant system. Free Radic Biol Med 2014; 66:75–87 [View Article] [PubMed]
    [Google Scholar]
  44. Bouillaut L, Self WT, Sonenshein AL. Proline-dependent regulation of Clostridium difficile Stickland metabolism. J Bacteriol 2013; 195:844–854 [View Article] [PubMed]
    [Google Scholar]
  45. Gencic S, Grahame DA, Metcalf WW. Diverse energy-conserving pathways in Clostridium difficile: growth in the absence of amino acid stickland acceptors and the role of the wood-ljungdahl pathway. J Bacteriol 2020; 202:20 [View Article]
    [Google Scholar]
  46. Pi H, Helmann JD. Sequential induction of Fur-regulated genes in response to iron limitation in Bacillus subtilis. Proc Natl Acad Sci U S A 2017; 114:12785–12790 [View Article] [PubMed]
    [Google Scholar]
  47. Zackular JP, Knippel RJ, Lopez CA, Beavers WN, Maxwell CN et al. ZupT facilitates Clostridioides difficile resistance to host-mediated nutritional immunity. mSphere 2020; 5:e00061-20. [View Article] [PubMed]
    [Google Scholar]
  48. Berges M, Michel A-M, Lassek C, Nuss AM, Beckstette M et al. Iron regulation in Clostridioides difficile. Front Microbiol 2018; 9:3183. [View Article] [PubMed]
    [Google Scholar]
  49. Ho TD, Ellermeier CD. Ferric uptake regulator fur control of putative iron acquisition systems in Clostridium difficile. J Bacteriol 2015; 197:2930–2940 [View Article] [PubMed]
    [Google Scholar]
  50. Cassat JE, Skaar EP. Iron in infection and immunity. Cell Host Microbe 2013; 13:509–519 [View Article] [PubMed]
    [Google Scholar]
  51. Winterbourn CC. Toxicity of iron and hydrogen peroxide: the Fenton reaction. Toxicol Lett 1995; 82–83:969–974 [View Article] [PubMed]
    [Google Scholar]
  52. Stevenson B, Wyckoff EE, Payne SM. Vibrio cholerae FeoA, FeoB, and FeoC interact to form a complex. J Bacteriol 2016; 198:1160–1170 [View Article] [PubMed]
    [Google Scholar]
  53. Cartron ML, Maddocks S, Gillingham P, Craven CJ, Andrews SC. Feo--transport of ferrous iron into bacteria. Biometals 2006; 19:143–157 [View Article] [PubMed]
    [Google Scholar]
  54. Dashper SG, Butler CA, Lissel JP, Paolini RA, Hoffmann B et al. A novel Porphyromonas gingivalis FeoB plays a role in manganese accumulation. J Biol Chem 2005; 280:28095–28102 [View Article] [PubMed]
    [Google Scholar]
  55. Roberts AP, Allan E, Mullany P. The impact of horizontal gene transfer on the biology of Clostridium difficile. Adv Microb Physiol 2014; 65:63–82 [View Article] [PubMed]
    [Google Scholar]
  56. Edwards AN, Krall EG, McBride SM. Strain-Dependent RstA regulation of Clostridioides difficile toxin production and sporulation. J Bacteriol 2020; 202:e00586-19. [View Article] [PubMed]
    [Google Scholar]
  57. Chen KY, Rathod J, Chiu YC, Chen JW, Tsai PJ et al. The transcriptional regulator Lrp contributes to toxin expression, sporulation, and swimming motility in Clostridium difficile. Front Cell Infect Microbiol 2019; 9:356. [View Article] [PubMed]
    [Google Scholar]
  58. Fortier LC. Bacteriophages contribute to shaping Clostridioides (Clostridium) difficile Species. Front Microbiol 2018; 9:2033. [View Article] [PubMed]
    [Google Scholar]
  59. Nakamura S, Nakashio S, Yamakawa K, Tanabe N, Nishida S. Carbohydrate fermentation by Clostridium difficile. Microbiol Immunol 1982; 26:107–111 [View Article] [PubMed]
    [Google Scholar]
  60. Neumann-Schaal M, Jahn D, Schmidt-Hohagen K. Metabolism the difficile way: the key to the success of the pathogen Clostridioides difficile. Front Microbiol 2019; 10:219. [View Article] [PubMed]
    [Google Scholar]
  61. Jackson S, Calos M, Myers A, Self WT. Analysis of proline reduction in the nosocomial pathogen Clostridium difficile. J Bacteriol 2006; 188:8487–8495 [View Article] [PubMed]
    [Google Scholar]
  62. Battaglioli EJ, Hale VL, Chen J, Jeraldo P, Ruiz-Mojica C et al. Clostridioides difficile uses amino acids associated with gut microbial dysbiosis in a subset of patients with diarrhea. Sci Transl Med 2018; 10:464 [View Article]
    [Google Scholar]
  63. Keeley TP, Mann GE. Defining physiological normoxia for improved translation of cell physiology to animal models and humans. Physiol Rev 2019; 99:161–234 [View Article] [PubMed]
    [Google Scholar]
  64. Knippel RJ, Zackular JP, Moore JL, Celis AI, Weiss A et al. Heme sensing and detoxification by HatRT contributes to pathogenesis during Clostridium difficile infection. PLoS Pathog 2018; 14:12e1007486 [View Article] [PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/mgen/10.1099/mgen.0.000738
Loading
/content/journal/mgen/10.1099/mgen.0.000738
Loading

Data & Media loading...

Supplements

Supplementary material 1

EXCEL

Supplementary material 2

EXCEL

Supplementary material 3

EXCEL

Supplementary material 4

EXCEL

Supplementary material 5

EXCEL

Supplementary material 6

PDF

Most cited this month Most Cited RSS feed

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