Exploiting the natural poly(3-hydroxyalkanoates) production capacity of Antarctic Pseudomonas strains: from unique phenotypes to novel biopolymers


Extreme environments are a unique source of microorganisms encoding metabolic capacities that remain largely unexplored. In this work, we isolated two Antarctic bacterial strains able to produce poly(3-hydroxyalkanoates) (PHAs), which were classified after 16S rRNA analysis as Pseudomonas sp. MPC5 and MPC6. The MPC6 strain presented nearly the same specific growth rate whether subjected to a temperature of 4 °C 0.18 (1/h) or 30 °C 0.2 (1/h) on glycerol. Both Pseudomonas strains produced high levels of PHAs and exopolysaccharides from glycerol at 4 °C and 30 °C in batch cultures, an attribute that has not been previously described for bacteria of this genus. The MPC5 strain produced the distinctive medium-chain-length-PHA whereas Pseudomonas sp. MPC6 synthesized a novel polyoxoester composed of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate-co-3-hydroxyoctanoate-co-3-hydroxydecanoate-co-3-hydroxydodecanoate). Batch bioreactor production of PHAs in MPC6 resulted in a titer of 2.6 (g/L) and 1.3 (g/L), accumulating 47.3% and 34.5% of the cell dry mass as PHA, at 30 and 4 °C, respectively. This study paves the way for using Antarctic Pseudomonas strains for biosynthesizing novel PHAs from low-cost substrates such as glycerol and the possibility to carry out the bioconversion process for biopolymer synthesis without the need for temperature control.

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  1. 1.

    Alteri CJ, Mobley HLT (2012) Escherichia coli physiology and metabolism dictates adaptation to diverse host microenvironments. Curr Opin Microbiol 15:3–9. https://doi.org/10.1016/j.mib.2011.12.004

    Article  CAS  PubMed  Google Scholar 

  2. 2.

    Angilletta M, Steury T, Sears M (2004) Temperature, growth rate, and body size in ectotherms: fitting pieces of a life-history puzzle. Integr Comp Biol 44:498–509

    Article  PubMed  Google Scholar 

  3. 3.

    Ayub ND, Pettinari MJ, Ruiz JA, López NI (2004) A polyhydroxybutyrate-producing Pseudomonas sp. isolated from antarctic environments with high stress resistance. Curr Microbiol 49:170–174. https://doi.org/10.1007/s00284-004-4254-2

    Article  CAS  Google Scholar 

  4. 4.

    Ayub ND, Tribelli PM, López NI (2009) Polyhydroxyalkanoates are essential for maintenance of redox state in the Antarctic bacterium Pseudomonas sp. 14-3 during low temperature adaptation. Extremophiles 13:59–66. https://doi.org/10.1007/s00792-008-0197-z

    Article  CAS  Google Scholar 

  5. 5.

    Bajerski F, Wagner D, Mangelsdorf K (2017) Cell membrane fatty acid composition of Chryseobacterium frigidisoli PB4(T), isolated from antarctic glacier forefield soils, in response to changing temperature and pH conditions. Front Microbiol 8:677. https://doi.org/10.3389/fmicb.2017.00677

    Article  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Bassas-Galia M, Nogales B, Arias S, Rohde M, Timmis KN, Molinari G (2012) Plant original Massilia isolates producing polyhydroxybutyrate, including one exhibiting high yields from glycerol. J Appl Microbiol 112:443–454. https://doi.org/10.1111/j.1365-2672.2011.05228.x

    Article  CAS  PubMed  Google Scholar 

  7. 7.

    Borrero-de Acuña JM, Aravena-Carrasco C, Gutierrez-Urrutia I, Duchens D, Poblete-Castro I (2019) Enhanced synthesis of medium-chain-length poly(3-hydroxyalkanoates) by inactivating the tricarboxylate transport system of Pseudomonas putida KT2440 and process development using waste vegetable oil. Process Biochem 77:23–30. https://doi.org/10.1016/j.procbio.2018.10.012

    Article  CAS  Google Scholar 

  8. 8.

    Borrero-de Acuña JM, Hidalgo-Dumont C, Pacheco N, Cabrera A, Poblete-Castro I (2017) A novel programmable lysozyme-based lysis system in Pseudomonas putida for biopolymer production. Sci Rep 7:4373. https://doi.org/10.1038/s41598-017-04741-2

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Cavicchioli R (2016) On the concept of a psychrophile. ISME J 10:793–795. https://doi.org/10.1038/ismej.2015.160

    Article  PubMed  Google Scholar 

  10. 10.

    Cheema S, Bassas-Galia M, Sarma PM, Lal B, Arias S (2012) Exploiting metagenomic diversity for novel polyhydroxyalkanoate synthases: production of a terpolymer poly(3-hydroxybutyrate-co-3-hydroxyhexanoate-co-3-hydroxyoctanoate) with a recombinant Pseudomonas putida strain. Bioresour Technol 103:322–328. https://doi.org/10.1016/j.biortech.2011.09.098

    Article  CAS  PubMed  Google Scholar 

  11. 11.

    Chen G-Q (2009) A microbial polyhydroxyalkanoates (PHA) based bio- and materials industry. Chem Soc Rev 38:2434–2446. https://doi.org/10.1039/b812677c

    Article  CAS  Google Scholar 

  12. 12.

    Ciesielski S, Górniak D, Możejko J, Świątecki A, Grzesiak J, Zdanowski M (2014) The diversity of bacteria isolated from Antarctic freshwater reservoirs possessing the ability to produce polyhydroxyalkanoates. Curr Microbiol 69:594–603. https://doi.org/10.1007/s00284-014-0629-1

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    D’Amico S, Collins T, Marx J-C, Feller G, Gerday C (2006) Psychrophilic microorganisms: challenges for life. EMBO Rep 7:385–389. https://doi.org/10.1038/sj.embor.7400662

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. 14.

    De Maayer P, Anderson D, Cary C, Cowan DA (2014) Some like it cold: understanding the survival strategies of psychrophiles. EMBO Rep 15:508–517

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. 15.

    Doi Y, Kitamura S, Abe H (1995) Microbial synthesis and characterization of poly(3-hydroxybutyrate-co-3-hydroxyhexanoate). Macromolecules 28:4822–4828. https://doi.org/10.1021/ma00118a007

    Article  CAS  Google Scholar 

  16. 16.

    Edgar RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 32:1792–1797. https://doi.org/10.1093/nar/gkh340

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. 17.

    Ferre-Guell A, Winterburn J (2018) Biosynthesis and characterization of polyhydroxyalkanoates with controlled composition and microstructure. Biomacromolecules 19:996–1005. https://doi.org/10.1021/acs.biomac.7b01788

    Article  CAS  PubMed  Google Scholar 

  18. 18.

    Fondi M, Maida I, Perrin E, Mellera A, Mocali S, Parrilli E, Tutino ML, Liò P, Fani R (2014) Genome-scale metabolic reconstruction and constraint-based modelling of the Antarctic bacterium Pseudoalteromonas haloplanktis TAC125. Environ Microbiol 17:751–766. https://doi.org/10.1111/1462-2920.12513

    Article  CAS  PubMed  Google Scholar 

  19. 19.

    Fong N, Burgess M, Barrow K, Glenn D (2001) Carotenoid accumulation in the psychrotrophic bacterium Arthrobacter agilis in response to thermal and salt stress. Appl Microbiol Biotechnol 56:750–756. https://doi.org/10.1007/s002530100739

    Article  CAS  PubMed  Google Scholar 

  20. 20.

    Frank S, Schmidt F, Klockgether J, Davenport CF, Gesell Salazar M, Völker U, Tümmler B (2011) Functional genomics of the initial phase of cold adaptation of Pseudomonas putida KT2440. FEMS Microbiol Lett 318:47–54

    Article  CAS  PubMed  Google Scholar 

  21. 21.

    Goh YS, Tan IKP (2012) Polyhydroxyalkanoate production by antarctic soil bacteria isolated from Casey Station and Signy Island. Microbiol Res 167:211–219. https://doi.org/10.1016/j.micres.2011.08.002

    Article  CAS  PubMed  Google Scholar 

  22. 22.

    Hartmann R, Hany R, Geiger T, Egli T, Witholt B, Zinn M (2004) Tailored biosynthesis of olefinic medium-chain-length poly[(R)-3-hydroxyalkanoates] in Pseudomonas putida GPo1 with improved thermal properties. Macromolecules 37:6780–6785

    Article  CAS  Google Scholar 

  23. 23.

    Higuchi-Takeuchi M, Morisaki K, Toyooka K, Numata K (2016) Synthesis of high-molecular-weight polyhydroxyalkanoates by marine photosynthetic purple bacteria. PLoS One 11:e0160981

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Jiao Y, Cody GD, Harding AK, Wilmes P, Schrenk M, Wheeler KE, Banfield JF, Thelen MP (2010) Characterization of extracellular polymeric substances from acidophilic microbial biofilms. Appl Environ Microbiol 76:2916–2922

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. 25.

    Lageveen RG, Huisman GW, Preusting H, Ketelaar P, Eggink G, Witholt B (1988) Formation of polyesters by Pseudomonas oleovorans: effect of substrates on formation and composition of poly-(R)-3-hydroxyalkanoates and poly-(R)-3-hydroxyalkenoates. Appl Environ Microbiol 54:2924–2932

    CAS  PubMed  PubMed Central  Google Scholar 

  26. 26.

    Li Z, Lin H, Ishii N, Chen G-Q, Inoue Y (2007) Study of enzymatic degradation of microbial copolyesters consisting of 3-hydroxybutyrate and medium-chain-length 3-hydroxyalkanoates. Polym Degrad Stab 92:1708–1714. https://doi.org/10.1016/j.polymdegradstab.2007.06.001

    Article  CAS  Google Scholar 

  27. 27.

    Lipson DA (2015) The complex relationship between microbial growth rate and yield and its implications for ecosystem processes. Front Microbiol 6:615. https://doi.org/10.3389/fmicb.2015.00615

    Article  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Madison LL, Huisman GW (1999) Metabolic engineering of poly(3-hydroxyalkanoates): from DNA to plastic. Microbiol Mol Biol Rev 63:21–53

    CAS  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Mansilla MC, Cybulski LE, Albanesi D, de Mendoza D (2004) Control of membrane lipid fluidity by molecular thermosensors. J Bacteriol 186:6681–6688. https://doi.org/10.1128/jb.186.20.6681-6688.2004

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Mocali S, Chiellini C, Fabiani A, Decuzzi S, de Pascale D, Parrilli E, Tutino ML, Perrin E, Bosi E, Fondi M, Lo Giudice A, Fani R (2017) Ecology of cold environments: new insights of bacterial metabolic adaptation through an integrated genomic–phenomic approach. Sci Rep 7:839. https://doi.org/10.1038/s41598-017-00876-4

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. 31.

    Morita RY (1975) Psychrophilic bacteria. Bacteriol Rev 39:144–167

    CAS  PubMed  PubMed Central  Google Scholar 

  32. 32.

    Nedwell DB (2006) Effect of low temperature on microbial growth: lowered affinity for substrates limits growth at low temperature. FEMS Microbiol Ecol 30:101–111. https://doi.org/10.1111/j.1574-6941.1999.tb00639.x

    Article  Google Scholar 

  33. 33.

    Nichols CM, Bowman JP, Guezennec J (2005) Effects of incubation temperature on growth and production of exopolysaccharides by an Antarctic Sea ice bacterium grown in batch culture. Appl Environ Microbiol 71:3519–3523. https://doi.org/10.1128/aem.71.7.3519-3523.2005

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Nikel PI, Chavarría M, Danchin A, de Lorenzo V (2016) From dirt to industrial applications: Pseudomonas putida as a synthetic biology chassis for hosting harsh biochemical reactions. Curr Opin Chem Biol 34:20–29. https://doi.org/10.1016/j.cbpa.2016.05.011

    Article  CAS  PubMed  Google Scholar 

  35. 35.

    Noghabi KA, Zahiri HS, Yoon SC (2007) The production of a cold-induced extracellular biopolymer by Pseudomonas fluorescens BM07 under various growth conditions and its role in heavy metals absorption. Process Biochem 42:847–855. https://doi.org/10.1016/j.procbio.2007.02.004

    Article  CAS  Google Scholar 

  36. 36.

    Obruca S, Sedlacek P, Koller M, Kucera D, Pernicova I (2017) Involvement of polyhydroxyalkanoates in stress resistance of microbial cells: biotechnological consequences and applications. Biotechnol Adv. https://doi.org/10.1016/j.biotechadv.2017.12.006

    Article  PubMed  Google Scholar 

  37. 37.

    Obruca S, Sedlacek P, Krzyzanek V, Mravec F, Hrubanova K, Samek O, Kucera D, Benesova P, Marova I (2016) Accumulation of poly(3-hydroxybutyrate) helps bacterial cells to survive freezing. PLoS One 11:e0157778–e0157778. https://doi.org/10.1371/journal.pone.0157778

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Oliva-Arancibia B, Órdenes-Aenishanslins N, Bruna N, Ibarra PS, Zacconi FC, Pérez-Donoso JM, Poblete-Castro I (2017) Co-synthesis of medium-chain-length polyhydroxyalkanoates and CdS quantum dots nanoparticles in Pseudomonas putida KT2440. J Biotechnol. https://doi.org/10.1016/j.jbiotec.2017.10.013

    Article  PubMed  Google Scholar 

  39. 39.

    Poblete-Castro I, Becker J, Dohnt K, Dos Santos VM, Wittmann C (2012) Industrial biotechnology of Pseudomonas putida and related species. Appl Microbiol Biotechnol 93:2279–2290. https://doi.org/10.1007/s00253-012-3928-0

    Article  CAS  PubMed  Google Scholar 

  40. 40.

    Poblete-Castro I, Binger D, Oehlert R, Rohde M (2014) Comparison of mcl-Poly(3-hydroxyalkanoates) synthesis by different Pseudomonas putida strains from crude glycerol: citrate accumulates at high titer under PHA-producing conditions. BMC Biotechnol 14:962

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Poblete-Castro I, Binger D, Rodrigues A, Becker J, Martins Dos Santos VAP, Wittmann C (2013) In-silico- driven metabolic engineering of Pseudomonas putida for enhanced production of poly-hydroxyalkanoates. Metab Eng 15:113–123

    Article  CAS  PubMed  Google Scholar 

  42. 42.

    Poblete-Castro I, Borrero-de Acuña JM, Nikel PI, Kohlstedt M, Wittmann C (2017) Host organism: Pseudomonas putida. In: Wittmann C, Liao JC (eds) Industrial biotechnology. Wiley-VCH, Weinheim, pp 299–326

    Google Scholar 

  43. 43.

    Poblete-Castro I, Wittmann C, Nikel PI (2019) Biochemistry, genetics and biotechnology of glycerol utilization in Pseudomonas species. Microb Biotechnol. https://doi.org/10.1111/1751-7915.13400

    Article  PubMed  Google Scholar 

  44. 44.

    Price PB, Sowers T (2004) Temperature dependence of metabolic rates for microbial growth, maintenance, and survival. Proc Natl Acad Sci USA 101:4631–4636. https://doi.org/10.1073/pnas.0400522101

    Article  CAS  PubMed  Google Scholar 

  45. 45.

    Prieto A, Escapa IF, Martínez V, Dinjaski N, Herencias C, de la Peña F, Tarazona N, Revelles O (2015) A holistic view of polyhydroxyalkanoate metabolism in Pseudomonas putida. Environ Microbiol 18:341–357

    Article  CAS  PubMed  Google Scholar 

  46. 46.

    Rehm BHA, Mitsky TA, Steinbüchel A (2001) Role of fatty acid de novo biosynthesis in polyhydroxyalkanoic acid (PHA) and rhamnolipid synthesis by Pseudomonads: establishment of the transacylase (PhaG)-mediated pathway for PHA biosynthesis in Escherichia coli. Appl Environ Microbiol. https://doi.org/10.1128/aem.67.7.3102-3109.2001

    Article  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Renner LD, Weibel DB (2011) Physicochemical regulation of biofilm formation. MRS Bull 36:347–355. https://doi.org/10.1557/mrs.2011.65

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Rodrigues DF, Tiedje JM (2008) Coping with our cold planet. Appl Environ Microbiol 74:1677–1686. https://doi.org/10.1128/aem.02000-07

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. 49.

    Sathiyanarayanan G, Bhatia SK, Song H-S, Jeon J-M, Kim J, Lee YK, Kim Y-G, Yang Y-H (2017) Production and characterization of medium-chain-length polyhydroxyalkanoate copolymer from Arctic psychrotrophic bacterium Pseudomonas sp. PAMC 28620. Int J Biol Macromol 97:710–720. https://doi.org/10.1016/j.ijbiomac.2017.01.053

    Article  CAS  PubMed  Google Scholar 

  50. 50.

    Seufferheld MJ, Alvarez HM, Farias ME (2008) Role of polyphosphates in microbial adaptation to extreme environments. Appl Environ Microbiol 74:5867–5874. https://doi.org/10.1128/aem.00501-08

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. 51.

    Shehata TE, Marr AG (1975) Effect of temperature on the size of Escherichia coli cells. J Bacteriol 124:857–862

    CAS  PubMed  PubMed Central  Google Scholar 

  52. 52.

    Stamatakis A (2014) RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics 30:1312–1313. https://doi.org/10.1093/bioinformatics/btu033

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. 53.

    Suutari M, Laakso S (1994) Microbial fatty acids and thermal adaptation. Crit Rev Microbiol 20:285–328. https://doi.org/10.3109/10408419409113560

    Article  CAS  PubMed  Google Scholar 

  54. 54.

    Tappel RC, Pan W, Bergey NS, Wang Q, Patterson IL, Ozumba OA, Matsumoto K, Taguchi S, Nomura CT (2014) Engineering Escherichia coli for improved production of short-chain-length-co-medium-chain-length poly[(R)-3-hydroxyalkanoate] (SCL-co-MCL PHA) copolymers from renewable nonfatty acid feedstocks. ACS Sustain Chem Eng 2:1879–1887. https://doi.org/10.1021/sc500217p

    Article  CAS  Google Scholar 

  55. 55.

    Tribelli PM, López NI (2011) Poly(3-hydroxybutyrate) influences biofilm formation and motility in the novel Antarctic species Pseudomonas extremaustralis under cold conditions. Extremophiles 15:541. https://doi.org/10.1007/s00792-011-0384-1

    Article  CAS  PubMed  Google Scholar 

  56. 56.

    Tripathi L, Wu L-P, Chen J, Chen G-Q (2012) Synthesis of Diblock copolymer poly-3-hydroxybutyrate-block-poly-3-hydroxyhexanoate [PHB-b-PHHx] by a β-oxidation weakened Pseudomonas putida KT2442. Microb Cell Fact 11:44. https://doi.org/10.1186/1475-2859-11-44

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. 57.

    Tsuge T (2016) Fundamental factors determining the molecular weight of polyhydroxyalkanoate during biosynthesis. Polym J 48:1051

    Article  CAS  Google Scholar 

  58. 58.

    Varas M, Valdivieso C, Mauriaca C, Ortíz-Severín J, Paradela A, Poblete-Castro I, Cabrera R, Chávez FP (2017) Multi-level evaluation of Escherichia coli polyphosphate related mutants using global transcriptomic, proteomic and phenomic analyses. Biochim Biophys Acta Gen Subj 1861:871–883. https://doi.org/10.1016/j.bbagen.2017.01.007

    Article  CAS  PubMed  Google Scholar 

  59. 59.

    Yang DC, Blair KM, Salama NR (2016) Staying in shape: the impact of cell shape on bacterial survival in diverse environments. Microbiol Mol Biol Rev 80:187–203

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. 60.

    Yergeau E, Michel C, Tremblay J, Niemi A, King TL, Wyglinski J, Lee K, Greer CW (2017) Metagenomic survey of the taxonomic and functional microbial communities of seawater and sea ice from the Canadian Arctic. Sci Rep 7:42242

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. 61.

    Yoneyama F, Yamamoto M, Hashimoto W, Murata K (2015) Production of polyhydroxybutyrate and alginate from glycerol by Azotobacter vinelandii under nitrogen-free conditions. Bioengineered 6:209–217. https://doi.org/10.1080/21655979.2015.1040209

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. 62.

    Zinn M, Witholt B, Egli T (2001) Occurrence, synthesis and medical application of bacterial polyhydroxyalkanoate. Adv Drug Deliv Rev 53:5–21. https://doi.org/10.1016/S0169-409X(01)00218-6

    Article  CAS  PubMed  Google Scholar 

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Nicolas Pacheco acknowledges the scholarship provided by Doctorado en Biotecnologia (UNAB). We thank the technical assistant of the Centro de Instrumentación of the Pontificia Universidad Católica de Chile through CONICYT-FONDEQUIP EQM120021. Andrés Marcoleta acknowledges to Macarena Varas for its aid in sample collection during the 53rd Chilean Antarctic Scientific Expedition, and to Johanna Rojas Salgado and José Ignacio Costa for their assistance in the isolation of the strains and its initial microbiological characterization.


This study was funded by CONICYT (Fondecyt Inicio Grant number 11150174) to I.P–C, and from the Chilean Antarctic Institute (INACH) through the Grant (RT_51-16) to A.E.M.

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Pacheco, N., Orellana-Saez, M., Pepczynska, M. et al. Exploiting the natural poly(3-hydroxyalkanoates) production capacity of Antarctic Pseudomonas strains: from unique phenotypes to novel biopolymers. J Ind Microbiol Biotechnol 46, 1139–1153 (2019). https://doi.org/10.1007/s10295-019-02186-2

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  • Antarctic Pseudomonas
  • Poly(3-hydroxyalkanoates)
  • Low temperature
  • Glycerol
  • Exopolysaccharide
  • Psychrophiles