Microalgae as bioreactors for bioplastic production

<p>Abstract</p> <p>Background</p> <p>Poly-3-hydroxybutyrate (PHB) is a polyester with thermoplastic properties that is naturally occurring and produced by such bacteria as <it>Ralstonia eutropha </it>H16 and <it>Bacillus megaterium</it>. In contrast to currently utilized plastics and most synthetic polymers, PHB is biodegradable, and its production is not dependent on fossil resources making this bioplastic interesting for various industrial applications.</p> <p>Results</p> <p>In this study, we report on introducing the bacterial PHB pathway of <it>R. eutropha </it>H16 into the diatom <it>Phaeodactylum tricornutum</it>, thereby demonstrating for the first time that PHB production is feasible in a microalgal system. Expression of the bacterial enzymes was sufficient to result in PHB levels of up to 10.6% of algal dry weight. The bioplastic accumulated in granule-like structures in the cytosol of the cells, as shown by light and electron microscopy.</p> <p>Conclusions</p> <p>Our studies demonstrate the great potential of microalgae like the diatom <it>P. tricornutum </it>to serve as solar-powered expression factories and reveal great advantages compared to plant based production systems.</p

Impact of Multiple β-Ketothiolase Deletion Mutations in Ralstonia eutropha H16 on the Composition of 3-Mercaptopropionic Acid-Containing Copolymers▿ §

β-Ketothiolases catalyze the first step of poly(3-hydroxybutyrate) [poly(3HB)] synthesis in bacteria by condensing two molecules of acetyl coenzyme A (acetyl-CoA) to acetoacetyl-CoA. Analyses of the genome sequence of Ralstonia eutropha H16 revealed 15 isoenzymes of PhaA in this bacterium. In this study, we generated knockout mutants of various phaA homologues to investigate their role in and contributions to poly(3HB) metabolism and to suppress biosynthesis of 3HB-CoA for obtaining enhanced molar 3-mercaptopriopionate (3MP) contents in poly(3HB-co-3MP) copolymers when cells were grown on gluconate plus 3-mercaptopropionate or 3,3′-dithiodipropionate. In silico sequence analysis of PhaA homologues, transcriptome data, and other aspects recommended the homologues phaA, bktB, H16_A1713/H16_B1771, H16_A1528, H16_B1369, H16_B0381, and H16_A0170 for further analysis. Single- and multiple-deletion mutants were generated to investigate the influence of these β-ketothiolases on growth and polymer accumulation. The deletion of single genes resulted in no significant differences from the wild type regarding growth and polymer accumulation during cultivation on gluconate or gluconate plus 3MP. Deletion of phaA plus bktB (H16Δ2 mutant) resulted in approximately 30% less polymer accumulation than in the wild type. Deletion of H16_A1713/H16_B1771, H16_A1528, H16_B0381, and H16_B1369 in addition to phaA and bktB gave no differences in comparison to the H16Δ2 mutant. In contrast, deletion of H16_A0170 additionally to phaA and bktB yielded a mutant which accumulated about 30% poly(3HB) (wt/wt of the cell dry weight [CDW]). Although we were not able to suppress poly(3HB) biosynthesis completely, the copolymer compositions could be altered significantly with a lowered percentage ratio of 3HB constituents (from 85 to 52 mol%) and an increased percentage ratio of 3MP constituents (from 15 to 48 mol%), respectively. In this study, we demonstrated that PhaA, BktB, and H16_A0170 are majorly involved in poly(3HB) synthesis in R. eutropha H16. A fourth β-ketothiolase or a combination of several of the other β-ketothiolases contributed to a maximum of only 30% (wt/wt of CDW) of the remaining (co)polymer

Analysis of the endophytic lifestyle and plant growth promotion of Burkholderia terricola ZR2-12

Members of the genus Burkholderia are highly versatile bacteria that can be beneficial as well as pathogenic for their eukaryotic hosts. Furthermore, many strains exhibit a remarkable biotechnological potential. To study the ecosystem function and lifestyle of B. terricola, we analysed the interactions with plants and survival in soil as well as the mechanisms behind it. We used a combination of in vitro and ad planta assays to study Burkholderia-plant interaction and assess the role of poly-beta-hydroxybutyrate (PHB). Additionally, DsRed-labelled bacteria were analysed by confocal laser scanning microscopy (CLSM) to study root colonisation. B. terricola ZR2-12 treatment resulted in enhanced growth of sugar beet plants with a more than doubled biomass relative to the non-treated control. The strain was a remarkable good root coloniser, which was found in rhizosphere as well as endorhiza of sugar beet up to 10 log(10) CFU g(-1). Using CLSM, we observed that ZR2-12 cells form large micro-colonies along the apoplastic spaces of the root. Xylem vessels were colonised by smaller aggregates and single cells, whereas in root tips mainly single cells were present. The colonisation patterns differed strongly between older and younger parts of the roots. PHB production of ZR2-12 (up to 70% (w/w) of cell dry mass) provided a competitive advantage for rhizosphere colonisation. B. terricola ZR2-12 belongs to the plant-associated Burkholderia cluster with biotechnological potential due to its excellent root colonisation and plant growth promotion