Conversion of Lignocellulosic Feedstocks into Bioethanol Using Extremophiles

  • Sean Michael Scully
  • Johann OrlygssonEmail author


  1. 1.

    Advantages of using thermophiles for bioethanol production

  2. 2.

    Considerations for using thermoanaerobes for processing lignocellulosic biomass

  3. 3.

    Utilization of combined bioprocesses such as SSF and CBP for bioethanol production

  4. 4.

    Recent advances in genetically modified thermoanaerobes for bioethanol production



Thermophilic anaerobes Second-generation biofuel Genetic engineering Consolidated processes Lignocellulose 


  1. Abdel-Banat BMA, Hosida H, Ano A, Nonklang S, Akada R (2010) High-temperature fermentation: how can processes for ethanol production at high temperatures become superior to the traditional process using mesophilic yeast? Appl Microbiol Biotechnol 85:861–867CrossRefPubMedGoogle Scholar
  2. Ahring BK, Jensen K, Nielsen P, Bjerre AB, Schmidt AS (1996) Pretreatment of wheat straw and conversion of xylose and xylan to ethanol by thermophilic anaerobic bacteria. Bioresour Technol 58:107–113CrossRefGoogle Scholar
  3. Ahring BK, Licht D, Schmidt AS, Sommer P, Thomsen AB (1999) Production of ethanol from wet oxidized wheat straw by Thermoanaerobacter mathranii. Bioresour Technol 68:3–9CrossRefGoogle Scholar
  4. Almarsdottir R, Sigurbjornsdottir MA, Orlygsson J (2012) Effects of various factors on ethanol yields from lignocellulosic biomass by Thermoanaerobacterium AK17. Biotechnol Bioeng 109:686–694CrossRefPubMedGoogle Scholar
  5. Andersen RL, Jensen KM, Mikkelsen MJ (2015) Continuous ethanol fermentation of pretreated lignocellulosic biomasses, waste biomass, molasses and syrup using the anaerobic, thermophilic bacterium Thermoanaerobacter italicus pentocrobe 411. PLoS ONE 10:e0136060CrossRefPubMedPubMedCentralGoogle Scholar
  6. Argyros DA, Tripathi SA, Barrett TF, Rogers SR, Feinberg LF, Olson DG, Foden JM, Miller BB, Lynd LR, Hogsett DA, Caiazza NC (2011) High ethanol titers from cellulose by using metabolically engineered thermophilic, anaerobic microbes. Appl Environ Microbiol 77:8288–8294CrossRefPubMedPubMedCentralGoogle Scholar
  7. Avci A, Donmez S (2006) Effect of zinc on ethanol production by two Thermoanaerobacter strains. Process Biochem 41:984–989CrossRefGoogle Scholar
  8. Balat M, Balat H, Öz C (2008) Progress in bioethanol processing. Prog Energy Combust Sci 34:551–573CrossRefGoogle Scholar
  9. Baskaran S, Ahn HJ, Lynd LR (1995) Investigation of the ethanol tolerance of Clostridium thermosaccharolyticum in continuous culture. Biotechnol Progress 11:276–281CrossRefGoogle Scholar
  10. Bhalla A, Bansal N, Kumar S et al (2013) Improved lignocellulose conversion to biofuels with thermophilic bacteria and thermostable enzymes. Bioresour Technol 128:751–759CrossRefPubMedGoogle Scholar
  11. Biswas R, Prabhu S, Lynd LR, Guss AM (2014) Increase in ethanol yield via elimination of lactate production in an ethanol-tolerant mutant of Clostridium thermocellum. PLoS ONE 9:e86389. Scholar
  12. Brynjarsdottir H, Wawiernia B, Orlygsson J (2012) Ethanol production from sugars and complex biomass by Thermoanaerobacter AK5: the effect of electron-scavenging systems on end-product formation. Energ Fuel 26:4568–4574CrossRefGoogle Scholar
  13. Chang T, Yao S (2011) Thermophilic, lignocellulolytic bacteria for ethanol production: current state and perspectives. Appl Microbiol Biotechnol 92:13–27CrossRefPubMedGoogle Scholar
  14. Crespo RE, Badshah M, Alvarez MT, Mattiason B (2012) Ethanol production by continuous fermentation of d-(+)-cellobiose, d-(+)-xylose and sugarcane bagasse hydrolysate using the thermoanaerobe Caloramator boliviensis. Bioresour Technol 103:186–191CrossRefPubMedGoogle Scholar
  15. Cripps RE, Eley K, Leak DJ, Rudd B, Taylor M, Todd M, Boakes S, Martin S, Atkinson T (2009) Metabolic engineering of Geobacillus thermoglucosidasius for high yield ethanol production. Metabol Eng 11:398–408CrossRefGoogle Scholar
  16. Desai SG, Guerinot ML, Lynd LR (2004) Cloning of l-lactate dehydrogenase and elimination of lactic acid production via gene knockout in Thermoanaerobacterium saccharolyticum JW/SL-YS485. Appl Microbiol Biotechnol 65:600–605CrossRefPubMedGoogle Scholar
  17. Elleuche S, Schröder C, Sahm K, Antranikian G (2014) Extremozymes—biocatalysts with unique properties from extremophilic microorganisms. Curr Opin Biotechnol 29:116–123CrossRefPubMedGoogle Scholar
  18. Georgieva TI, Ahring BK (2007) Evaluation of continuous ethanol fermentation of dilute-acid corn stover hydrolysate using thermophilic anaerobic bacterium Thermoanaerobacter BG1L1. Appl Microbiol Biotechnol 77:61–68CrossRefPubMedGoogle Scholar
  19. Georgieva TI, Mikkelsen MJ, Ahring BK (2008) Ethanol production from wet-exploded wheat straw hydrolysate by thermophilic anaerobic bacterium Thermoanaerobacter BG1L1 in a continuous immobilized reactor. Appl Biochem Biotechnol 145:99–110CrossRefPubMedGoogle Scholar
  20. He Q, Lokken PM, Chen S et al (2009) Characterization of the impact of acetate and lactate on ethanolic fermentation by Thermoanaerobacter ethanolicus. Bioresour Technol 100:5955–5965CrossRefPubMedGoogle Scholar
  21. Jeffries TW (2006) Engineering yeasts for xylose metabolism. Curr Opin Biotechnol 17:320–326CrossRefPubMedGoogle Scholar
  22. Jessen JE, Orlygsson J (2012) Production of ethanol from sugars and lignocellulosic biomass by Thermoanaerobacter J1 isolated from a hot spring in Iceland. J Biomed Biotechnol 2012:1. Scholar
  23. Lacis LS, Lawford HG (1988) Ethanol-production from xylose by Thermoanaerobacter ethanolicus in batch and continuous culture. Arch Microbiol 150:48–55CrossRefGoogle Scholar
  24. Lin L, Tu O, Huang R, Teng L, Zeng X, Song H, Wang K, Zhou Q, Li Y, Cui Q, He Z, Zhou J, Xu J (2013) Microevolution from shock to adaptation revealed strategies improving ethanol tolerance and production in Thermoanaerobacter. Biotechnol Biofuels 6:103CrossRefPubMedPubMedCentralGoogle Scholar
  25. Lovitt RW, Longin R, Zeikus JG (1984) Ethanol production by thermophilic bacteria: physiological comparison of solvent effects on parent and alcohol-tolerant strains of Clostridium thermohydrosulfuricum. Appl Environ Microbiol 48:171–177PubMedPubMedCentralGoogle Scholar
  26. Lovitt RW, Shen GJ, Zeikus JG (1988) Ethanol production by thermophilic bacteria: biochemical basis for ethanol and hydrogen tolerance in Clostridium thermohydrosulfuricum. J Bacteriol 170:2809–2815CrossRefPubMedPubMedCentralGoogle Scholar
  27. Lynd LR (1996) Overview and evaluation of fuel ethanol from cellulosic biomass: technology, economics, the environment, and policy. Annu Rev Energy Environ 21:403–465CrossRefGoogle Scholar
  28. Lynd LR, Weimer PJ, van Zyl WH, Pretorius IS (2002) Microbial cellulose utilization fundamentals and biotechnology. Microbiol Mol Biol Rev 66:506–577CrossRefPubMedPubMedCentralGoogle Scholar
  29. Lynd LR, van Zyl WH, McBride JE, Laser M (2005) Consolidated bioprocessing of cellulosic biomass: an update. Curr Opin Biotechnol 16:577–583CrossRefPubMedGoogle Scholar
  30. Mai V, Wiegel J (2000) Advances in development of genetic system for Thermoanaerobacterium spp.: expression of genes encoding hydrolytic enzymes, development of second shuttle vector, and integration of genes into the chromosome. Appl Environ Microbiol 66:4817–4821CrossRefPubMedPubMedCentralGoogle Scholar
  31. Mai V, Lorenz WW, Wiegel J (1997) Transformation of Thermoanaerobacterium sp. strain JW/SL-YS485 with plasmid pIKM1 conferring kanamycin resistance. FEMS Microbiol Lett 148:163–167CrossRefGoogle Scholar
  32. Menon V, Rao M (2012) Trends in bioconversion of lignocellulose: biofuels, platform chemicals & biorefinery concept. Prog Energy Combust Sci 38:522–550CrossRefGoogle Scholar
  33. Mosier N, Wyman C, Dale B, Elander R, Lee YY, Holtzapple M, Ladisch M (2005) Features of promising technologies for pretreatment of lignocellulosic biomass. Bioresour Technol 96:673–686CrossRefPubMedGoogle Scholar
  34. Orlygsson J (2012) Ethanol production from biomass by a moderate thermophile, Clostridium AK1. Icel Agric Sci 25:25–35Google Scholar
  35. Ostergaard S, Olsson L, Nielsen J (2000) Metabolic engineering of Saccharomyces cerevisiae. Microbiol Mol Biol Rev 64:34–50CrossRefPubMedPubMedCentralGoogle Scholar
  36. Pereira FB, Gomes DG, Guimares PMR, Teixera JA, Domingues L (2012) Cell recycling during repeated very high gravity bio-ethanol fermentations using the industrial Saccharomyces cerevisiae strain PE-2. Biotechnol Lett 34:45–53CrossRefPubMedGoogle Scholar
  37. Rani KS, Swamy MV, Seenayya G (1997) Increased ethanol production by metabolic modulation of cellulose fermentation in Clostridium thermocellum. Biotechnol Lett 19:819–823CrossRefGoogle Scholar
  38. RFA - Renewable Fuels Association (2013) World fuel ethanol production. Available online: Accessed 30 Sept 2014
  39. Rogers PL, Lee KJ, de Tribe DE (1979) Kinetics of alcohol production by Zymomonas mobilis at high sugar concentrations. Biotechnol Lett 1:165–170CrossRefGoogle Scholar
  40. Sánchez ÓJ, Cardona CA (2008) Trends in biotechnological production of fuel ethanol from different feedstocks. Bioresour Technol 99:5270–5295CrossRefPubMedGoogle Scholar
  41. Scully SM, Orlygsson J (2015) Recent advances in second generation ethanol production by thermophilic bacteria. Energies 8:1–30CrossRefGoogle Scholar
  42. Shao X, Raman B, Zhu MJ, Mielenz JR, Brown SD, Guss AM, Lynd LR (2011) Mutant selection and phenotypic and genetic characterization of ethanol-tolerant strains of Clostridium thermocellum. Appl Microbiol Biotechnol 92:641–652CrossRefPubMedGoogle Scholar
  43. Shaw AJ, Podkaminer KK, Desai SG, Bardslev JS, Rogers SR, Thorne PG, Hogsett DA, Lynd LL (2008) Metabolic engineering of a thermophilic bacterium to produce ethanol at high yield. Proc Natl Acad Sci U S A 105:13769–13774CrossRefPubMedPubMedCentralGoogle Scholar
  44. Shaw AJ, Hogsett DA, Lynd LR (2009) Identification of the [FeFe]-hydrogenase responsible for hydrogen generation in Thermoanaerobacterium saccharolyticum and demonstration of increased ethanol yield via hydrogenase knockout. J Bacteriol 191:6457–6464CrossRefPubMedPubMedCentralGoogle Scholar
  45. Shaw AJ, Hogsett DA, Lynd LR (2010) Natural competence in Thermoanaerobacter and Thermoanaerobacterium species. Appl Environ Microbiol 76:4713–4719CrossRefPubMedPubMedCentralGoogle Scholar
  46. Sittijunda S, Tomas AF, Reungsang A, O-Thong S, Angelidaki I (2013) Ethanol production from glucose and xylose by immobilized Thermoanaerobacter pentosaceus at 70 °C in an up-flow anaerobic sludge blanket (UASB) reactor. Bioresour Technol 143:598–607CrossRefPubMedGoogle Scholar
  47. Sommer P, Georgieva T, Ahring BK (2004) Potential for using thermophilic anaerobic bacteria for bioethanol production from hemicellulose. Biochem Soc Trans 32:283–289CrossRefPubMedGoogle Scholar
  48. Sveinsdottir M, Baldursson SRB, Orlygsson J (2009) Ethanol Production from monosugars and lignocellulosic biomass by thermophilic bacteria isolated from Icelandic hot springs. Iceland Agric Sci 22:45–58Google Scholar
  49. Talebnia F, Karakashev D, Angelidaki I (2010) Production of bioethanol from wheat straw: an overview on pretreatment, hydrolysis and fermentation. Bioresour Technol 101:4744–4753CrossRefPubMedGoogle Scholar
  50. Tan H-T, Corbin KR, Fincher GB (2016) Emerging technologies for the production of renewable liquid transport fuels from biomass sources enriched in plant cell walls. Front Plant Sci 7:1854PubMedPubMedCentralGoogle Scholar
  51. Taylor MP, Eley KL, Martin S, Tuffin MI, Burton SG, Cowan DA (2009) Thermophilic ethanologenesis: future prospects for second-generation bioethanol production. Trends Biotechnol 27:398–405CrossRefPubMedGoogle Scholar
  52. Timmons MD, Knutson BL, Nokes SE, Strobel HJ (2009) Analysis of composition and structure of Clostridium thermocellum membranes from wild-type and ethanol-adapted strains. Appl Microbiol Biotechnol 82:929–939CrossRefPubMedGoogle Scholar
  53. Tomás AF (2013) Optimization of bioethanol production from carbohydrate rich wastes by extreme thermophilic microorganisms. Ph.D. Thesis, DTU, Copenhagen, DenmarkGoogle Scholar
  54. Tomas AF, Karakashev D, Angelidaki I (2011) Effect of xylose and nutrients concentration on ethanol production by a newly isolated extreme thermophilic bacterium. Water Res Technol 64:341–347CrossRefGoogle Scholar
  55. Tomas AF, Karakashef D, Angelidaki I (2013) Thermoanaerobacter pentosaceus sp. nov., an anaerobic, extremely thermophilic, high ethanol-yielding bacterium isolated from household waste. Int J Syst Evol Microbiol 63:2396–2404CrossRefPubMedGoogle Scholar
  56. Tripathi SA, Olson DG, Argyros DA, Miller BB, Barrett TF, Murphy DM, McCool JD, Warner AK, Raigarhia VB, Lynd LR, Hogsett DA, Caiazza NC (2010) Development of pyrF-based genetic system for targeted gene deletion in Clostridium thermocellum and creation of a pta mutant. Appl Environ Microbiol 76:6591–6599CrossRefPubMedPubMedCentralGoogle Scholar
  57. Turner P, Mamo G, Karlson EN (2007) Potential and utilization of thermophiles and thermostable enzymes in biorefining. Microb Cell Factor 6:9CrossRefGoogle Scholar
  58. Tyurin MV, Lynd LR, Wiegel J (2006) 13 Gene transfer systems for obligately anaerobic thermophilic bacteria. In: Rainey FA, Oren A (eds) Methods in microbiology, vol 35. Academic Press Ltd/Elsevier Science Ltd, London, pp 309–330Google Scholar
  59. Van Zyl LJ, Taylor MP, Eley K, Tuffin M, Cowan DA (2014) Engineering pyruvate decarboxylase-mediated ethanol production in the thermophilic host Geobacillus thermoglucosidasius. Appl Microbiol Biotechnol 98:1247–1259CrossRefPubMedGoogle Scholar
  60. Viponik Z, Jessen JE, Scully SM, Orlygsson J (2016) Effect of culture conditions on hydrogen production by Thermoanaerobacter strain AK68. Int J Hydrog Energ 41:181–189CrossRefGoogle Scholar
  61. Vohra M, Manwar J, Manmode R, Padgilwar S, Patil S (2014) Bioethanol production: feedstock and current technologies. J Environ Chem Eng 2:573–584CrossRefGoogle Scholar
  62. Wiegel J, Ljungdahl LG (1981) Thermoanaerobacter ethanolicus gen. Nov., spec. Nov., a new, extreme thermophilic, anaerobic bacterium. Arch Microbiol 128:343–348CrossRefGoogle Scholar
  63. Wiegel J, Carreira LH, Mothershed CP, Puls J (1983) Production of ethanol from bio-polymers by anaerobic, thermophilic, and extreme thermophilic bacteria. II. Thermoanaerobacter ethanolicus JW200 and its mutants in batch cultures and resting cell experiments. Biotechnol Bioeng 13:193–205Google Scholar
  64. Xu L, Tschirner U (2014) Immobilized anaerobic fermentation for bio-fuel production by Clostridium co-culture. Bioprocess Biosyst Eng 37:1551–1559CrossRefPubMedGoogle Scholar
  65. Xu Q, Singh A, Himmel ME (2009) Perspectives and new directions for the production of bioethanol using consolidated bioprocessing of lignocellulose. Curr Opin Biotechnol 20:364–371CrossRefPubMedGoogle Scholar
  66. Yao S, Mikkelsen MJ (2010a) Metabolic engineering to improve ethanol production in Thermoanaerobacter mathranii. Appl Microbiol Biotechnol 88:199–208CrossRefPubMedGoogle Scholar
  67. Yao S, Mikkelsen MJ (2010b) Identification and overexpression of a bifunctional aldehyde/alcohol dehydrogenase responsible for ethanol production in Thermoanaerobacter mathranii. J Mol Microbiol Biotechnol 19:123–133CrossRefPubMedGoogle Scholar

Copyright information

© Springer International Publishing AG, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Faculty of Natural Resource SciencesUniversity of AkureyriAkureyriIceland

Personalised recommendations