Skip to main content
SpringerLink
Log in

Dual stress factors adaptive evolution for high EPA production in Schizochytrium sp. and metabolomics mechanism analysis

  • Research Paper
  • Published:
Bioprocess and Biosystems Engineering Aims and scope Submit manuscript

Abstract

Eicosapentaenoic acid (EPA) is a vital ɷ-3 polyunsaturated fatty acid (PUFA) for human body with various physiological functions. In this study, we proposed an adaptive evolutionary strategy based on high-temperature and high-oxygen two-factor stress to increase the EPA production capacity of Schizochytrium. High-temperature stress was used to increase EPA yield, and high oxygen was implemented to continuously stimulate cell growth and lipid accumulation. The biomass and EPA production of ALE-D50 reached 35.33 g/L and 1.54 g/L, which were 43.85% and 71.11% higher than that of the original strain, respectively. Lower in vivo reactive oxygen species levels indicated that the evolved strain possessed stronger antioxidant activity. Liquid chromatography–mass spectrometry metabolomics showed that enhanced glucose consumption and glycolysis metabolism, as well as a weakened tricarboxylic acid cycle and reduced amino acid metabolic tributaries in the evolved strain, might be associated with increased growth and EPA synthesis. Finally, the lipid production and EPA production in a fed-batch fermentation were further increased to 48.93 g/L and 3.55 g/L, improving by 54.30% and 90.86%, respectively. This study provides a novel pathway for promoting EPA biosynthesis in Schizochytrium.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Data availability

The datasets generated during the current study are available from the corresponding author on reasonable request.

References

  1. Šimat V, Vlahović J, Soldo B, Generalić Mekinić I, Čagalj M, Hamed I, Skroza D (2020) Production and characterization of crude oils from seafood processing by-products. Food Biosci 33:1–8. https://doi.org/10.1016/j.fbio.2019.100484

    Article  CAS  Google Scholar 

  2. Sherratt SCR, Libby P, Dawoud H, Bhatt DL, Malinski T, Mason RP (2023) Eicosapentaenoic acid (EPA) reduces pulmonary endothelial dysfunction and inflammation due to changes in protein expression during exposure to particulate matter air pollution. Biomed Pharmacother 162:1–12. https://doi.org/10.1016/j.biopha.2023.114629

    Article  CAS  Google Scholar 

  3. Izumi T, Nanaura H, Iguchi N, Ozaki M, Sugie K (2020) Low serum eicosapentaenoic acid levels in cryptogenic stroke with active cancer. J Stroke Cerebrovasc 29(8):1–8. https://doi.org/10.1016/j.jstrokecerebrovasdis.2020.104892

    Article  Google Scholar 

  4. Kumari A, Pabbi S, Tyagi A (2023) Recent advances in enhancing the production of long chain omega-3 fatty acids in microalgae. Crit Rev Food Sci Nutr. https://doi.org/10.1080/10408398.2023.2226720

    Article  PubMed  Google Scholar 

  5. Das UN (2021) Bioactive lipids in COVID-19-further evidence. Archs Med Res 52(1):107–120. https://doi.org/10.1016/j.arcmed.2020.09.006

    Article  CAS  Google Scholar 

  6. Barta DG, Coman V, Vodnar DC (2021) Microalgae as sources of omega-3 polyunsaturated fatty acids: Biotechnological aspects. Algal Res 58:1–13. https://doi.org/10.1016/j.algal.2021.102410

    Article  Google Scholar 

  7. Wei X, Wang Y, Liu X, Hu Z, Qian J, Shi T, Ye C (2023) Metabolic analysis of Schizochytrium sp mutants with high EPA content achieved with ARTP mutagenesis screening. Bioproc Biosyst Eng 46(6):893–901. https://doi.org/10.1007/s00449-023-02874-5

    Article  CAS  Google Scholar 

  8. Huang J, Aki T, Hachida K, Yokochi T, Kawamoto S, Shigeta S, Suzuki O (2001) Profile of polyunsaturated fatty acids produced by Thraustochytrium sp KK17–3. J Am Oil Chem Soc 78(6):605–610. https://doi.org/10.1007/s11746-001-0312-1

    Article  CAS  Google Scholar 

  9. Ling X, Zhou H, Yang Q, Yu S, Li J, Li Z, Lu Y (2020) Functions of enyolreductase (ER) domains of PKS cluster in lipid synthesis and enhancement of PUFAs accumulation in Schizochytrium limacinum SR21 using triclosan as a regulator of ER. Microorganisms 8(2):1–17. https://doi.org/10.3390/microorganisms8020300

    Article  CAS  Google Scholar 

  10. Ren LJ, Chen SL, Geng LJ, Ji XJ, Xu X, Song P, Huang H (2018) Exploring the function of acyltransferase and domain replacement in order to change the polyunsaturated fatty acid profile of Schizochytrium sp. Algal Res 29:193–201. https://doi.org/10.1016/j.algal.2017.11.021

    Article  Google Scholar 

  11. Wang J, Wang Y, Wu Y, Fan Y, Zhu C, Fu X, Mou H (2021) Application of microalgal stress responses in industrial microalgal production systems. Mar Drugs 20(1):1–15. https://doi.org/10.3390/md20010030

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Zhang B, Wu J, Meng F (2021) Adaptive Laboratory evolution of microalgae: a review of the regulation of growth, stress resistance, metabolic processes, and biodegradation of pollutants. Front Microbiol 12:1–8. https://doi.org/10.3389/fmicb.2021.737248

    Article  Google Scholar 

  13. Qi F, Zhang M, Chen Y, Jiang X, Lin J, Cao X, Huang J (2017) A lignocellulosic hydrolysate-tolerant Aurantiochytrium sp. mutant strain for docosahexaenoic acid production. Bioresource Technol 227:221–226. https://doi.org/10.1016/j.biortech.2016.12.011

    Article  CAS  Google Scholar 

  14. Li X, Pei G, Liu L, Chen L, Zhang W (2017) Metabolomic analysis and lipid accumulation in a glucose-tolerant Crypthecodinium cohnii strain obtained by adaptive laboratory evolution. Bioresource Technol 235:87–95. https://doi.org/10.1016/j.biortech.2017.03.049

    Article  CAS  Google Scholar 

  15. Hu X, Tang X, Bi Z, Zhao Q, Ren L (2021) Adaptive evolution of microalgae Schizochytrium sp. under high temperature for efficient production of docosahexaeonic acid. Algal Res 54:1–9. https://doi.org/10.1016/j.algal.2021.102212

    Article  Google Scholar 

  16. Sun XM, Ren LJ, Ji XJ, Chen SL, Guo DS, Huang H (2016) Adaptive evolution of Schizochytrium sp. by continuous high oxygen stimulations to enhance docosahexaenoic acid synthesis. Bioresource Technol 211:374–381. https://doi.org/10.1016/j.biortech.2016.03.093

    Article  CAS  Google Scholar 

  17. Sun XM, Ren LJ, Bi ZQ, Ji XJ, Zhao QY, Jiang L, Huang H (2018) Development of a cooperative two-factor adaptive-evolution method to enhance lipid production and prevent lipid peroxidation in Schizochytrium sp. Biotechnol Biofuels 11(65):1–16. https://doi.org/10.1186/s13068-018-1065-4

    Article  CAS  Google Scholar 

  18. Zhang R, Luo Y, Yu X, Dang M, Hu X, Ren L (2023) Adaptive evolution of Schizochytrium sp. under light and H2O2 condition to regulate its fatty acid and terpene biosynthesis. Algal Res 72:1–8. https://doi.org/10.1016/j.algal.2023.103127

    Article  Google Scholar 

  19. Zhong H, Zhang M, Chen L, Liu W, Tao Y (2023) Development of Schizochytrium sp. strain HS01 with high-DHA and low-saturated fatty acids production by multi-pronged adaptive evolution. Biotechnol Lett:1–11. https://doi.org/10.1007/s10529-023-03378-8.

  20. Ou Y, Li Y, Feng S, Wang Q, Yang H (2023) Transcriptome analysis reveals an eicosapentaenoic acid accumulation mechanism in a Schizochytrium sp mutant. Microbiol Spectr 11(3):1–15. https://doi.org/10.1128/spectrum.00130-23

    Article  CAS  Google Scholar 

  21. Zhao B, Li Y, Mbifile MD, Li C, Yang H, Wang W (2017) Improvement of docosahexaenoic acid fermentation from Schizochytrium sp AB-610 by staged pH control based on cell morphological changes. Eng Life Sci 17(9):981–988. https://doi.org/10.1002/elsc.201600249

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Chang G, Gao N, Tian G, Wu Q, Chang M, Wang X (2013) Improvement of docosahexaenoic acid production on glycerol by Schizochytrium sp. S31 with constantly high oxygen transfer coefficient. Bioresource Technol 142:400–406. https://doi.org/10.1016/j.biortech.2013.04.107

    Article  CAS  Google Scholar 

  23. Bao Z, Zhu Y, Zhang K, Feng Y, Chen X, Lei M, Yu L (2021) High-value utilization of the waste hydrolysate of Dioscorea zingiberensis for docosahexaenoic acid production in Schizochytrium sp. Bioresource Technol 336:1–8. https://doi.org/10.1016/j.biortech.2021.125305

    Article  CAS  Google Scholar 

  24. Liu ZX, You S, Tang BP, Wang B, Sheng S, Wu FA, Wang J (2019) Inositol as a new enhancer for improving lipid production and accumulation in Schizochytrium sp SR21. Environ Sci Pollut R 26(35):35497–35508. https://doi.org/10.1007/s11356-019-06056-3

    Article  CAS  Google Scholar 

  25. Pahlavan M, Jalili H, Noroozi M, Moradi Y, Saba F (2019) Optimization of culture conditions for growth of the Aurantiochytrium sp shy, isolated from the Persian Gulf. Iran J Fish Sci 18(4):656–671. https://doi.org/10.22092/ijfs.2018.117491

    Article  Google Scholar 

  26. Silva DDV, Dussán KJ, Hernández V, Silva SSd, Cardona CA, Felipe MdGdA (2016) Effect of volumetric oxygen transfer coefficient (kLa) on ethanol production performance by Scheffersomyces stipitis on hemicellulosic sugarcane bagasse hydrolysate. Biochem Eng J 112:249–257. https://doi.org/10.1016/j.bej.2016.04.012

    Article  CAS  Google Scholar 

  27. Jakobsen AN, Aasen IM, Josefsen KD, Strom AR (2008) Accumulation of docosahexaenoic acid-rich lipid in thraustochytrid Aurantiochytrium sp strain T66: effects of N and P starvation and O2 limitation. Appl Microbiol Biot 80(2):297–306. https://doi.org/10.1007/s00253-008-1537-8

    Article  CAS  Google Scholar 

  28. Bi ZQ, Ren LJ, Hu XC, Sun XM, Zhu SY, Ji XJ, Huang H (2018) Transcriptome and gene expression analysis of docosahexaenoic acid producer Schizochytrium sp. under different oxygen supply conditions. Biotechnol Biofuels 11:1–13. https://doi.org/10.1186/s13068-018-1250-5

    Article  CAS  Google Scholar 

  29. Hu F, Clevenger AL, Zheng P, Huang Q, Wang Z (2020) Low-temperature effects on docosahexaenoic acid biosynthesis in Schizochytrium sp. TIO01 and its proposed underlying mechanism. Biotechnol Biofuels 13:1–14. https://doi.org/10.1186/s13068-020-01811-y

    Article  CAS  Google Scholar 

  30. Xing G, Yuan H, Yang J, Li J, Gao Q, Li W, Wang E (2018) Integrated analyses of transcriptome, proteome and fatty acid profilings of the oleaginous microalga Auxenochlorella protothecoides UTEX 2341 reveal differential reprogramming of fatty acid metabolism in response to low and high temperatures. Algal Res 33:16–27. https://doi.org/10.1016/j.algal.2018.04.028

    Article  Google Scholar 

  31. Bao Z, Zhu Y, Feng Y, Zhang K, Zhang M, Wang Z, Yu L (2022) Enhancement of lipid accumulation and docosahexaenoic acid synthesis in Schizochytrium sp. H016 by exogenous supplementation of sesamol. Bioresource Technol 345:1–9. https://doi.org/10.1016/j.biortech.2021.126527

    Article  CAS  Google Scholar 

  32. Amaro HM, Fernandes F, Valentao P, Andrade PB, Sousa-Pinto I, Malcata FX, Guedes AC (2015) Effect of solvent system on extractability of lipidic components of Scenedesmus obliquus (M2–1) and Gloeothece sp on antioxidant scavenging capacity thereof. Mar Drugs 13(10):6453–6471. https://doi.org/10.3390/md13106453

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Mariam I, Kareya MS, Nesamma AA, Jutur PP (2021) Delineating metabolomic changes in native isolate Aurantiochytrium for production of docosahexaenoic acid in presence of varying carbon substrates. Algal Res 55:1–11. https://doi.org/10.1016/j.algal.2021.102285

    Article  Google Scholar 

  34. Li Z, Meng T, Hang W, Cao X, Ni H, Shi Y, He N (2021) Regulation of glucose and glycerol for production of docosahexaenoic acid in Schizochytrium limacinum SR21 with metabolomics analysis. Algal Res 58:1–10. https://doi.org/10.1016/j.algal.2021.102415

    Article  Google Scholar 

  35. Yu XJ, Sun J, Sun YQ, Zheng JY, Wang Z (2016) Metabolomics analysis of phytohormone gibberellin improving lipid and DHA accumulation in Aurantiochytrium sp. Biochem Eng J 112:258–268. https://doi.org/10.1016/j.bej.2016.05.002

    Article  CAS  Google Scholar 

  36. Chen Z, Zheng Z, Yi C, Wang F, Niu Y, Li H (2016) Intracellular metabolic changes in Saccharomyces cerevisiae and promotion of ethanol tolerance during the bioethanol fermentation process. RSC Adv 6(107):105046–105055. https://doi.org/10.1039/c6ra19254h

    Article  CAS  Google Scholar 

  37. Fina A, Millard P, Albiol J, Ferrer P, Heux S (2023) High throughput 13C-metabolic flux analysis of 3-hydroxypropionic acid producing Pichia pastoris reveals limited availability of acetyl-CoA and ATP due to tight control of the glycolytic flux. Microb Cell Fact 22(1):1–16. https://doi.org/10.1186/s12934-023-02123-0

    Article  CAS  Google Scholar 

  38. Yang J, Song X, Wang L, Cui Q (2020) Comprehensive analysis of metabolic alterations in Schizochytrium sp. strains with different DHA content. J Chromatogr B 1160:1–8. https://doi.org/10.1016/j.jchromb.2020.122193

    Article  CAS  Google Scholar 

  39. Cui GZ, Ma Z, Liu YJ, Feng Y, Sun Z, Cheng Y, Cui Q (2016) Overexpression of glucose-6-phosphate dehydrogenase enhanced the polyunsaturated fatty acid composition of Aurantiochytrium sp. SD116. Algal Res 19:138–145. https://doi.org/10.1016/j.algal.2016.08.005

    Article  Google Scholar 

  40. Yildizli A, Çevik S, Ünyayar S (2018) Effects of exogenous myo-inositol on leaf water status and oxidative stress of Capsicum annuum under drought stress. Acta Physiol Plant 40(6):1–10. https://doi.org/10.1007/s11738-018-2690-z

    Article  CAS  Google Scholar 

  41. Dong L, Wang F, Chen L, Zhang W (2023) Metabolomic analysis reveals the responses of docosahexaenoic-acid-producing Schizochytrium under hyposalinity conditions. Algal Res 70:1–10. https://doi.org/10.1016/j.algal.2023.102987

    Article  Google Scholar 

  42. Geng L, Chen S, Sun X, Hu X, Ji X, Huang H, Ren L (2019) Fermentation performance and metabolomic analysis of an engineered high-yield PUFA-producing strain of Schizochytrium sp. Bioproc Biosyst Eng 42(1):71–81. https://doi.org/10.1007/s00449-018-2015-z

    Article  CAS  Google Scholar 

  43. Wang S, Lan C, Wang Z, Wan W, Zhang H, Cui Q, Song X (2020) Optimizing eicosapentaenoic acid production by grafting a heterologous polyketide synthase pathway in the thraustochytrid Aurantiochytrium. J Agric Food Chem 68(40):11253–11260. https://doi.org/10.1021/acs.jafc.0c04299

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Key Research and Development Program of China (Grant No. 2022YFC3401300), and the National Natural Science Foundation of China (Grant No. 32371540, 21878128).

Author information

Authors and Affiliations

  1. The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, WuXi, 214122, People’s Republic of China

    Ying Ou, Yu Qin, Shoushuai Feng & Hailin Yang

Authors
  1. Ying Ou
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yu Qin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Shoushuai Feng
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hailin Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Ying Ou methodology, resources, formal analysis, investigation, and writing—original draft; Yu Qin formal analysis and investigation; Shoushuai Feng: conceptualization, funding acquisition, supervision, and writing—review & editing; Hailin Yang conceptualization, funding acquisition, supervision, and writing—review & editing.

Corresponding author

Correspondence to Hailin Yang.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (DOCX 1126 KB)

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ou, Y., Qin, Y., Feng, S. et al. Dual stress factors adaptive evolution for high EPA production in Schizochytrium sp. and metabolomics mechanism analysis. Bioprocess Biosyst Eng 47, 863–875 (2024). https://doi.org/10.1007/s00449-024-03013-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00449-024-03013-4

Keywords

Search

Navigation