Extremophilic microorganisms are considered being very promising candidates for biotechnological production of various products including polyhydroxyalkanoates (PHA). The aim of this work was to evaluate the PHA production potential of a novel PHA-producing thermophilic Gram-positive isolate Aneurinibacillus sp. H1. This organism was capable of efficient conversion of glycerol into poly(3-hydroxybutyrate) (P3HB), the homopolyester of 3-hydroxybutyrate (3HB). In flasks experiment, under optimal cultivation temperature of 45 °C, the P3HB content in biomass and P3HB titers reached 55.31% of cell dry mass and 2.03 g/L, respectively. Further, the isolate was capable of biosynthesis of PHA copolymers and terpolymers containing high molar fractions of 3-hydroxyvalerate (3HV) and 4-hydroxybutyrate (4HB). Especially 4HB contents in PHA were very high (up to 91 mol %) when 1,4-butanediol was used as a substrate. Based on these results, it can be stated that Aneurinibacillus sp. H1 is a very promising candidate for production of PHA with tailored material properties.

ARENA Arbeitsgemeinschaft für Ressourcenschonende and Nachhaltige Technologien Inffeldgasse 21b 8010 Graz Austria

Biology Centre The Czech Academy of Sciences v v i Branisovska 31 370 05 Ceske Budejovice Czech Republic

Faculty of Chemistry Brno University of Technology Purkynova 118 612 00 Brno Czech Republic

Faculty of Science University of South Bohemia Branisovska 31 370 05 Ceske Budejovice Czech Republic

Institute of Scientific Instruments of the Czech Academy of Sciences v v i Kralovopolska 147 612 64 Brno Czech Republic

Office of Research and Management c o Institute of Chemistry NAWI Graz University of Graz Heinrichstrasse 28 6 8010 Graz Austria

Zobrazit více v PubMed

Zeldes B.M., Keller M.W., Loder A.J., Straub C.T., Adams M.W.W., Kelly R.M. Extremely thermophilic microorganisms as metabolic engineering platforms for production of fuels and industrial chemicals. Front. Microbiol. 2015;6:1209. doi: 10.3389/fmicb.2015.01209. PubMed DOI PMC

Ranawat P., Rawat S. Stress response physiology of thermophiles. Arch. Microbiol. 2017;199:391–414. PubMed

Chen G.-Q., Jiang X.-R. Next generation industrial biotechnology based on extremophilic bacteria. Curr. Opin. Biotechnol. 2018;50:94–100. PubMed

Obruca S., Sedlacek P., Koller M., Kucera D., Pernicova I. Involvement of polyhydroxyalkanoates in stress resistance of microbial cells: Biotechnological consequences and applications. Biotechnol. Adv. 2018;36:856–870. doi: 10.1016/j.biotechadv.2017.12.006. PubMed DOI

Koller M., Maršálek L., Miranda de Sousa Dias M., Braunegg G. Producing microbial polyhydroxyalkanoate (PHA) biopolyesters in a sustainable manner. New Biotechnol. 2017;37:24–38. doi: 10.1016/j.nbt.2016.05.001. PubMed DOI

Sudesh K., Abe H., Doi Y. Synthesis, structure and properties of polyhydroxyalkanoates: Biological polyesters. Prog. Polym. Sci. 2000;25:1503–1555. doi: 10.1016/S0079-6700(00)00035-6. DOI

Kunioka M., Tamaki A., Doi Y. Crystalline and thermal properties of bacterial copolyesters: Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) and poly(3-hydroxybutyrate-co-4-hydroxybutyrate) Macromolecules. 1989;22:694–697. doi: 10.1021/ma00192a031. DOI

Koller M. Biodegradable and biocompatible polyhydroxy-alkanoates (PHA): Auspicious microbial macromolecules for pharmaceutical and therapeutic applications. Molecules. 2018;23:362. doi: 10.3390/molecules23020362. PubMed DOI PMC

Koller M. Chemical and biochemical engineering approaches in manufacturing Polyhydroxyalkanoate (PHA) biopolyesters of tailored structure with focus on the diversity of building blocks. Chem. Biochem. Eng. Q. 2019;32:413–438. doi: 10.15255/CABEQ.2018.1385. DOI

Koller M. Switching from petro-plastics to microbial polyhydroxyalkanoates (PHA): The biotechnological escape route of choice out of the plastic predicament? Eur. Biotechnol. J. 2019;3:32–44.

Dietrich K., Dumont M.-J., Del Rio L.F., Orsat V. Sustainable PHA production in integrated lignocellulose biorefineries. New Biotechnol. 2019;49:161–168. PubMed

Favaro L., Basaglia M., Casella S. Improving polyhydroxyalkanoate production from inexpensive carbon sources by genetic approaches: A review. Biofuel Bioprod. Biorefin. 2019;13:208–227. doi: 10.1002/bbb.1944. DOI

Ibrahim M.H., Willems A., Steinbüchel A. Isolation and characterization of new poly (3HB)-accumulating star-shaped cell aggregates-forming thermophilic bacteria. J. Appl. Microbiol. 2010;109:1579–1590. doi: 10.1111/j.1365-2672.2010.04786.x. PubMed DOI

Pantazaki A.A., Tambaka M.G., Langlois V., Guerin P., Kyriakidis D.A. Polyhydroxyalkanoate (PHA) biosynthesis in Thermus thermophilus: Purification and biochemical properties of PHA synthase. Mol. Cell. Biochem. 2003;254:173–183. doi: 10.1023/A:1027373100955. PubMed DOI

Sheu D.S., Chen W.M., Yang J.Y., Chang R.C. Thermophilic bacterium Caldimonas taiwanensis produces poly (3-hydroxybutyrate-co-3-hydroxyvalerate) from starch and valerate as carbon sources. Enzyme Microb. Technol. 2009;44:289–294.

Pernicova I., Novackova I., Sedlacek P., Kourilova X., Koller M., Obruca S. Application of osmotic challenge for enrichment of microbial consortia in polyhydroxyalkanoates producing thermophilic and thermotolerant bacteria and their subsequent isolation. Int. J. Biol. Macromol. 2020;144:698–704. PubMed

Nováková D., Švec P., Zeman M., Busse H.-J., Mašlaňová I., Pantůček R., Králová S., Krištofová L., Sedláček I. Pseudomonas leptonychotis sp. nov., isolated from weddell seals in Antarctica. Int. J. Syst. Evol. Microbiol. 2020;70:302–308. PubMed

Obruca S., Sedlacek P., Mravec F., Krzyzanek V., Nebesarova J., Samek O., Kucera D., Benesova P., Hrubanova K., Milerova M., et al. The presence of PHB granules in cytoplasm protects non-halophilic bacterial cells against the harmful impact of hypertonic environments. New Biotechnol. 2017;39:68–80. doi: 10.1016/j.nbt.2017.07.008. PubMed DOI

Obruca S., Benesova P., Oborna J., Marova I. Application of protease-hydrolyzed whey as a complex nitrogen source to increase poly(3-hydroxybutyrate) production from oils by Cupriavidus necator. Biotechnol. Lett. 2014;36:775–781. PubMed

Novackova I., Kucera D., Porizka J., Pernicova I., Sedlacek P., Koller M., Kovalcik A., Obruca S. Adaptation of Cupriavidus necator to levulinic acid for enhanced production of P(3HB-co-3HV) copolyesters. Biochem. Eng. J. 2019;151:107350.

Johnston B., Radecka I., Hill D., Chiellini E., Ilieva V.I., Sikorska W., Musioł M., Ziȩba M., Marek A.A., Keddie D., et al. The microbial production of Polyhydroxyalkanoates from waste polystyrene fragments attained using oxidative degradation. Polymers. 2018;10:957. doi: 10.3390/polym10090957. PubMed DOI PMC

Fadzil F.I.M., Mizuno S., Hiroe A., Nomura C.T., Tsuge T. Low Carbon concentration feeding improves medium-chain-length polyhydroxyalkanoate production in Escherichia coli strains with defective β-oxidation. Front. Bioeng. Biotechnol. 2018;6:178. doi: 10.3389/fbioe.2018.00178. PubMed DOI PMC

Pernicova I., Kucera D., Nebesarova J., Kalina M., Novackova I., Koller M., Obruca S. Production of polyhydroxyalkanoates on waste frying oil employing selected Halomonas strains. Bioresour. Technol. 2019;292:122028. PubMed

Ye J., Hu D., Yin J., Huang W., Xiang R., Zhang L., Wang X., Han J., Chen G.-Q. Stimulus response-based fine-tuning of polyhydroxyalkanoate pathway in Halomonas. Metab. Eng. 2020;57:85–95. doi: 10.1016/j.ymben.2019.10.007. PubMed DOI

Singh A.K., Srivastava J.K., Chandel A.K., Sharma L., Mallick N., Singh S.P. Biomedical applications of microbially engineered polyhydroxyalkanoates: An insight into recent advances, bottlenecks, and solutions. Appl. Microbiol. Biotechnol. 2019;103:2007–2032. PubMed

Kumar P., Patel S.K.S., Lee J.-K., Kalia V.C. Extending the limits of Bacillus for novel biotechnological applications. Biotechnol. Adv. 2013;31:1543–1561. doi: 10.1016/j.biotechadv.2013.08.007. PubMed DOI

Kumar P., Kim B.S. Valorization of polyhydroxyalkanoates production process by co-synthesis of value-added products. Bioresour. Technol. 2018;269:544–556. doi: 10.1016/j.biortech.2018.08.120. PubMed DOI

Kumar P., Ray S., Patel S.K.S., Lee J.-K., Kalia V.C. Bioconversion of crude glycerol to polyhydroxyalkanoate by Bacillus thuringiensis under non-limiting nitrogen conditions. Int. J. Biol. Macromol. 2015;78:9–16. doi: 10.1016/j.ijbiomac.2015.03.046. PubMed DOI

Shida O., Takagi H., Kadowaki K., Komagata K. Proposal for two new genera, Brevibacillus gen. nov. and Aneurinibacillus gen. nov. Int. J. Syst. Bacteriol. 1996;46:939–946. doi: 10.1099/00207713-46-4-939. PubMed DOI

Xiao Z., Zhang Y., Xi L., Huo F., Zhao J.-Y., Li J. Thermophilic production of polyhydroxyalkanoates by a novel Aneurinibacillus strain isolated from Gudao oilfield, China. J. Basic Microb. 2015;55:1125–1133. doi: 10.1002/jobm.201400843. PubMed DOI

Mravec F., Obruca S., Krzyzanek V., Sedlacek P., Hrubanova K., Samek O., Kucera D., Benesova P., Nebesarova J. Accumulation of PHA granules in Cupriavidus necator as seen by confocal fluorescence microscopy. FEMS Microbiol. Lett. 2016;363:fnw094. doi: 10.1093/femsle/fnw094. PubMed DOI

Kucera D., Pernicová I., Kovalcik A., Koller M., Mullerova L., Sedlacek P., Mravec F., Nebesarova J., Kalina M., Marova I., et al. Characterization of the promising poly(3-hydroxybutyrate) producing halophilic bacterium Halomonas halophila. Bioresour. Technol. 2018;256:552–556. doi: 10.1016/j.biortech.2018.02.062. PubMed DOI

Sadykov M.R., Ahn J.-S., Widhelm T.J., Eckrich V.M., Endres J.L., Driks A., Rutkowski G.E., Wingerd K.L., Bayles K.W. Poly(3-hydroxybutyrate) fuels the tricarboxylic acid cycle and de novo lipid biosynthesis during Bacillus anthracis sporulation. Mol. Microbiol. 2017;104:793–803. PubMed

Valappil S.P., Misra S.K., Boccaccini A.R., Keshavarz T., Bucke C., Roy I. Large-scale production and efficient recovery of PHB with desirable material properties, from the newly characterised Bacillus cereus SPV. J. Biotechnol. 2007;132:251–258. doi: 10.1016/j.jbiotec.2007.03.013. PubMed DOI

Ciriminna R., Pina C.D., Rossi M., Pagliaro M. Understanding the glycerol market. Eur. J. Lipid Sci. Technol. 2014;116:1432–1439.

Mohandas S.P., Balan L., Jayanath G., Anoop B.S., Philip R., Cubelio S.S., Bright Singh I.S. Biosynthesis and characterization of polyhydroxyalkanoate from marine Bacillus cereus MCCB 281 utilizing glycerol as carbon source. Int. J. Biol. Macromol. 2018;119:380–392. doi: 10.1016/j.ijbiomac.2018.07.044. PubMed DOI

Gahlawat G., Soni S.K. Valorization of waste glycerol for the production of poly (3-hydroxybutyrate) and poly (3-hydroxybutyrate-co-3-hydroxyvalerate) copolymer by Cupriavidus necator and extraction in a sustainable manner. Bioresour. Technol. 2017;243:492–501. doi: 10.1016/j.biortech.2017.06.139. PubMed DOI

Zain N.F.M., Abdullah W.N.W., Shun T.J., Keong L.C., Samian M.R. Optimization of polyhydroxyalkanoate (PHA) production by Burkholderia cepacia BPT1213 utilizing waste glycerol as the sole carbon source. Malaysian J. Microbiol. 2018;14:164–171.

Hermann-Krauss C., Koller M., Muhr A., Fasl H., Stelzer F., Braunegg G. Archaeal production of polyhydroxyalkanoate (PHA) co- and terpolyesters from biodiesel industry-derived by-products. Archaea. 2013;2013:129268. doi: 10.1155/2013/129268. PubMed DOI PMC

Hsiao L.-J., Lee M.-C., Chuang P.-J., Kuo Y.-Y., Lin J.-H., Wu T.-M., Li S.-Y. The production of poly(3-hydroxybutyrate) by thermophilic Caldimonas manganoxidans from glycerol. J. Polym. Res. 2018;25:85. doi: 10.1007/s10965-018-1486-6. DOI

Ibrahim M.H., Steinbüchel A. High-cell-density cyclic fed-batch fermentation of a poly (3-hydroxybutyrate)-accumulating thermophile, Chelatococcus sp. strain MW10. Appl. Environ. Microbiol. 2010;76:7890–7895. doi: 10.1128/AEM.01488-10. PubMed DOI PMC

Vigneswari S., Vijaya S., Majid M.I.A., Sudesh K., Sipaut C.S., Azizan M.N.M., Amirul A.A. Enhanced production of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) copolymer with manipulated variables and its properties. J. Ind. Microbiol. Biotechnol. 2009;36:547–556. doi: 10.1007/s10295-009-0525-z. PubMed DOI

Huong K.-H., Mohd Yahya A.R., Amirul A.A. Pronounced synergistic influence of mixed substrate cultivation on single step copolymer P(3HB-co-4HB) biosynthesis with a wide range of 4HB monomer composition. J. Chem. Technol. Biotechnol. 2014;89:1023–1029. doi: 10.1002/jctb.4195. DOI

Lee W.-H., Azizan M.N.M., Sudesh K. Efects of culture conditions on the composition of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) synthesized by Comamonas acidovorans. Polym. Degrad. Stab. 2004;84:129–134. doi: 10.1016/j.polymdegradstab.2003.10.003. DOI

Kucera D., Novackova I., Pernicova I., Sedlacek P., Obruca S. Biotechnological Production of poly(3-hydroxybutyrate-co-4-hydroxybutyrate-co-3-hydroxyvalerate) terpolymer by Cupriavidus sp. DSM 19379. Bioengineering. 2019;6:74. doi: 10.3390/bioengineering6030074. PubMed DOI PMC

Evaluating stress resilience of cyanobacteria through flow cytometry and fluorescent viability assessment

Degradation of P(3HB-co-4HB) Films in Simulated Body Fluids

Effects of Differing Monomer Compositions on Properties of P(3HB-co-4HB) Synthesized by Aneurinibacillus sp. H1 for Various Applications

Biotechnological Conversion of Grape Pomace to Poly(3-hydroxybutyrate) by Moderately Thermophilic Bacterium Tepidimonas taiwanensis