Isolation and characterization of polyhydroxyalkanoate producing bacteria from legume rhizosphere soils in Me Linh commune, Hanoi
Author affiliations
DOI:
https://doi.org/10.15625/vjbt-23144Keywords:
Bioplastic, polyhydroxyalkanoates (PHAs), Priestia spp., Pseudomonas spp., soil bacteriaAbstract
The increasing demand for sustainable bioplastics has driven the search for polyhydroxyalkanoate (PHA)-producing microorganisms from diverse ecological niches. In this study, we isolated and characterized native soil bacteria with the ability to accumulate PHAs from the rhizosphere of leguminous crops in Me Linh commune, Hanoi. A total of 206 bacterial isolates were obtained from eight soil samples and screened for intracellular PHA granules using Sudan Black B and Nile Blue A staining. Of these, 21 isolates tested positive for PHA accumulation, and five strains (ML53, ML71, ML91, ML113, and ML205) were selected for further analysis. Quantitative fermentation experiments revealed that strains ML113, ML91, and ML71 achieved the highest PHA contents, ranging from 20.7% to 24.8% of dry cell weight (DCW). Fourier-transform infrared (FTIR) spectroscopy confirmed the presence of functional groups characteristic of PHB and PHBV biopolymers. Morphological, biochemical, and molecular analyses—particularly 16S rRNA gene sequencing and phylogenetic tree construction—revealed that the selected isolates belong to three distinct genera: Pseudomonas (ML53, closely related to P. putida), Micrococcus (ML91, closely related to M. luteus), and Priestia (ML71 and ML205, clustered with P. megaterium, and ML113, closely related to P. aryabhattai). The results highlight the legume rhizosphere as a rich source of genetically diverse and metabolically competent PHA-producing bacteria with promising potential for future bioplastic development.
Downloads
References
Arcos-Hernández, M. V., Laycock, B., Donose, B. C., Pratt, S., Halley, P., Al-Luaibi, S., et al. (2013). Physicochemical and mechanical properties of mixed culture polyhydroxyalkanoate (PHBV). European Polymer Journal, 49(4), 904-913. https://doi.org/10.1016/j.eurpolymj.2012.10.025
Barillot, C. D. C., Sarde, C. O., Bert, V., Tarnaud, E., & Cochet, N. (2013). A standardized method for the sampling of rhizosphere and rhizoplan soil bacteria associated to a herbaceous root system. Annals of Microbiology, 63, 471–476. https://doi.org/10.1007/s13213-012-0491-y
Belal, E. B. (2013). Production of poly-β-hydroxybutyric acid (PHB) by Rhizobium elti and Pseudomonas stutzeri. Current Research Journal of Biological Sciences, 5(6), 273-284. http://dx.doi.org/10.19026/crjbs.5.5429
Chen, G. Q. (2010). Plastics completely synthesized by bacteria: Polyhydroxyalkanoates. In Plastics from Bacteria, 17–37. https://doi.org/10.1007/978-3-642-03287-5_2
Chen, J. Y., Song, G. & Chen, G. Q. (2006). A lower specificity PhaC2 synthase from Pseudomonas stutzeri catalyses the production of copolyesters consisting of short-chain-length and medium-chain-length 3-hydroxyalkanoates. Antonie Van Leeuwenhoek, 89, 157-167. https://doi.org/10.1007/s10482-005-9019-9
Claus, D., & Berkeley, R. C. W. (1986). Genus Bacillus Cohn 1872, in: Bergey’s Manual of Systematic Bacteriology, Vol. 2 (P. H. A. Sneath, ed.), Bergey’s Manual Trust, Williams and Wilkins, Baltimore, 1105–1139.
Getino, L., Martín, J. L., & Chamizo-Ampudia, A. (2024). A review of polyhydroxyalkanoates: Characterization, production, and application from waste. Microorganisms, 12(10), 2028. https://doi.org/10.3390/microorganisms12102028
Israni, N., & Shivakumar, S. (2015). Evaluation of Upstream Process parameters influencing the growth associated PHA accumulation in Bacillus sp. Ti3. Journal of Scientific & Industrial Research, 74, 290-295.
Joyline, M., & Aruna, K. (2019). Production and characterization of polyhydroxyalkanoates (PHA) by Bacillus megaterium strain JHA using inexpensive agro-industrial wastes. International Journal of Scientific Research, 10, 33359–33374. http://dx.doi.org/10.24327/ijrsr.2019.1007.3656
Koller, M., & Rodríguez-Contreras, A. (2015). Techniques for tracing PHA-producing organisms and for qualitative and quantitative analysis of intra- and extracellular PHA. Engineering in Life Sciences, 15, 558-581. https://doi.org/10.1002/elsc.201400228
Laycock, B., Halley, P., Pratt, S., Werker, A., & Lant, P. (2013). The chemomechanical properties of microbial polyhydroxyalkanoates. Progress in Polymer Science, 38(3–4), 536–583. https://doi.org/10.1016/j.progpolymsci.2012.06.003
Lee, S. Y. (1996). Bacterial polyhydroxyalkanoates. Biotechnology and Bioengineering, 49(1), 1–14. https://doi.org/10.1002/(SICI)1097-0290(19960105)49:1%3C1::AID-BIT1%3E3.0.CO;2-P
Li, M. L., Doudin, K., Robins, D. B., Tetradis-Mairis, G., Wong, T. S., Tee, K. L. (2025). Microbial synthesis of polyhydroxyalkanoate blends with engineered Pseudomonas putida, New Biotechnology, 88, 161-170. https://doi.org/10.1016/j.nbt.2025.05.004
Madigan, M. T., Bender, K. S., Buckley, D. H., Sattley, W. M., & Stahl, D. A. (2018). Brock Biology of Microorganisms (15th ed.). Pearson.
Musa, H., Bolanle, B. B., Kasim, F. H., Arbain, D. (2016). Screening and production of Polyhydroxybutyrate (PHB) by bacterial strains isolated from rhizosphere soil of groundnut plants. Sains Malaysiana, 45, 1469-1476.
Nishida, M., Tanaka, T., Hayakawa, Y., Nishida, M. (2018). Solid state nuclear magnetic resonance (NMR) and nuclear magnetic relaxation time analyses of molecular mobility and compatibility of plasticized polyhydroxyalkanoates (PHA) copolymers. Polymers, 10(5), 506. https://doi.org/10.3390/polym10050506
Orita, I., Iwazawa, R., Nakamura, S., & Fukui, T. (2012). Identification of mutation points in Cupriavidus necator NCIMB 11599 and genetic reconstitution of glucose-utilization ability in wild strain H16 for polyhydroxyalkanoate production. Journal of Bioscience and Bioengineering, 113(1), 63–69. https://doi.org/10.1016/j.jbiosc.2011.09.014
Ostle, A. G., Holt, J. G. (1982). Nile blue A as a fluorescent stain for poly-beta-hydroxybutyrate. Applied and Environmental Microbiology, 44(1), 238-241. https://journals.asm.org/doi/10.1128/aem.44.1.238-241.1982
Palleroni, N. J. (2010). The Pseudomonas story. Environmental Microbiology, 12(6), 1377–1383. https://doi.org/10.1111/j.1462-2920.2009.02041.x
Patil, T. D., Ghosh, S., Agarwal, A., Patel, S. K. S., Tripathi, A. D., Mahato, D. K., et al. (2024). Production, optimization, scale up and characterization of polyhydoxyalkanoates copolymers utilizing dairy processing waste. Scientific Reports, 14(1), 1620. https://doi.org/10.1038/s41598-024-52098-0
Râpă, M., Stefan, L. M., Seciu-Grama, A.-M., Gaspar-Pintiliescu, A., Matei, E., Zaharia, C., et al. (2022). Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (P(3HB-co-3HV))/Bacterial Cellulose (BC) biocomposites for potential use in biomedical applications. Polymers, 14(24), 5544. https://doi.org/10.3390/polym14245544
Rivera-Briso A. L. & Serrano-Aroca Á. (2018). Poly(3-Hydroxybutyrate-co-3-Hydroxyvalerate) Enhancement strategies for advanced applications. Polymers (Basel), 10(7), 732. https://doi.org/10.3390/polym10070732
Schlegel, H. G., Lafferty, R., & Krauss, I. (1970). The isolation of mutants not accumulating poly-β-hydroxybutyric acid. Archiv für Mikrobiologie, 71(3), 283–294. https://doi.org/10.1007/BF00410161
Shah, S., & Kumar, A. (2021). Production and characterization of polyhydroxyalkanoates from industrial waste using soil bacterial isolates. Brazilian Journal of Microbiology, 52(2), 715–726. https://doi.org/10.1007/s42770-021-00452-z
Spiekermann, P., Rehm, B. H., Kalscheuer, R., Baumeister, D., & Steinbüchel, A. (1999). A sensitive, viable-colony staining method using Nile red for direct screening of bacteria that accumulate polyhydroxyalkanoic acids and other lipid storage compounds. Archives of Microbiology, 171(2), 73–80. https://doi.org/10.1007/s002030050681
Trakunjae, C., Sudesh, K., Neoh, S. Z., Boondaeng, A., Apiwatanapiwat, W., Janchai, P., et al. (2022). Biosynthesis of P(3HB-co-3HHx) copolymers by a newly engineered strain of Cupriavidus necator PHB−4/pBBR_CnPro-phaCRp for skin tissue engineering application. Polymers, 14(19), 4074. https://doi.org/10.3390/polym14194074
Zhang, J., & Cran, M. J. J. (2022). Production of polyhydroxyalkanoate nanoparticles using a green solvent. Applied Polymer Science, 139(23), e52319. https://doi.org/10.1002/app.52319
Downloads
Published
How to Cite
Issue
Section
Funding data
-
Vietnam Academy of Science and Technology
Grant numbers TĐNSH0.07/22-24
