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      Producción de polihidroxialcanoatos por bacterias halófilas nativas utilizando almidón de cáscaras de Solanum tuberosum L Translated title: Production of polyhydroxyalkanoates by native halophilic bacteria using Solanum tuberosum L. shell starch

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          Abstract

          El objetivo del presente estudio fue determinar la concentración de almidón de cáscaras de Solanum tuberosum L. "papa" para la producción de polihidroxialcanoatos, PHA, por bacterias halófilas nativas, como una alternativa para disminuir los costos de producción de estos biopolímeros, posibles reemplazantes de los plásticos provenientes del petróleo. Las bacterias fueron aisladas de muestras de agua de ocho salinas y fueron enriquecidas en caldo HM1 a 30 °C, con 5, 10, 15, 20, 25 y 30 g 100 mL-1 de NaCl. A continuación, se realizaron diluciones, se tomaron alícuotas y se sembraron en agar HM1, obteniéndose 203 aislados de bacterias halófilas que fueron cultivados en caldo HM2 con 10 g.L-1 de glucosa como fuente de carbono y entre las que el 38,92% presentó gránulos de PHA teñidos por Sudan Negro B. Los 20 aislados con gránulos de PHA en el 65 95 % de las células nuevamente fueron llevadas a fermentación, alcanzando 0,174 0,889 g.g-1 de rendimiento Y (p/x). Se seleccionaron tres aislados con los mayores valores entre los que Halomonas sp. M4C1 desarrolló y sintetizó PHA en caldo HM2 con 5, 10, 15 y 20 g.L-1 de almidón como fuente de carbono, alcanzando 0,019; 0,016; 0,007 y 0,006 g.L-1 de PHA, con 0,177; 0,111; 0,056 y 0,066 g.L-1 de biomasa, después de 20, 40, 24 y 16 horas respectivamente. El mayor rendimiento de 0,144 g.g-1 le correspondió a 10 g.L-1 de almidón demostrándose que con esta concentración es factible la producción de PHA por las bacterias halófilas nativas

          Translated abstract

          The aim of this study was to determine the concentration of starch Solanum tuberosum L. "potato" peels for the production of polyhydroxyalkanoates, PHA, from native halophilic bacteria as an alternative to reduce production costs of these biopolymers, possible replacements for petrochemical plastics. The bacteria were isolated of water samples of eight saline Lambayeque region and were enriched in HM1 broth at 30 °C with 5, 10, 15, 20, 25 and 30 g 100 mL-1 NaCl. Dilutions were performed subsequently, aliquots were taken and plated on HM1 agar, 203 isolates of halophilic bacteria were obtained, they were grown in HM2 broth with 10 g.L-1 glucose as carbon source and the 38.92% of the isolates showed PHA granules stained by Sudan Black B. Twenty bacteria with PHA granules in 65 - 75% over carried to fermentation, reaching between 0.174 to 0.889 g.g-1 of yield Y (p / x). Three isolates were selected with the highest values among which Halomonas sp M4C1 grew and synthesized PHA in HM2 broth with 5, 10, 15 and 20 g.L-1 of starch as carbon source, reaching 0.019; 0.016; 0.007 y 0.006 g.L-1 of PHA, with 0.177; 0.111; 0.056 and 0.066 g.L-1 of biomass after 20, 40, 24 and 16 hours respectively. The highest yield of 0.144 g.g-1 corresponded to 10 g.L-1 of starch demonstrating that this concentration is feasible PHA production by native halophilic bacteria

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          Most cited references45

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          Production of polyhydroxyalkanoates from inexpensive extruded rice bran and starch by Haloferax mediterranei.

          Low-cost raw materials can be used to reduce significantly the production cost of polyhydroxyalkanoates (PHA). In this study, extruded rice bran (ERB) and extruded cornstarch (ECS) were used as carbon sources to produce PHA by an archaea, Haloferax mediterranei, which cannot use native rice bran or cornstarch as a carbon source. By employing pH-stat control strategy to maintain pH at 6.9-7.1 in a 5-liter jar fermentor using ERB:ECS (1:8 g/g) as the major carbon source, we obtained a cell concentration of 140 g/L, PHA concentration of 77.8 g/L and PHA content of 55.6 wt.% in a repeated fed-batch fermentation. In contrast, when ECS was used as the major carbon source, we obtained 62.6 g/L cell concentration, 24.2 g/L PHA concentration and 38.7 wt.% PHA content. Under a hyper-saline condition and with no nitrogen-limitation restriction, the repeated fed-batch process can be sustained a long time for the mass production of PHA.
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            Identification of polyhydroxyalkanoates in Halococcus and other haloarchaeal species

            Polyhydroxyalkanoates (PHAs) are accumulated in many prokaryotes. Several members of the Halobacteriaceae produce poly-3-hydroxybutyrate (PHB), but it is not known if this is a general property of the family. We evaluated identification methods for PHAs with 20 haloarchaeal species, three of them isolates from Permian salt. Staining with Sudan Black B, Nile Blue A, or Nile Red was applied to screen for the presence of PHAs. Transmission electron microscopy and 1H-nuclear magnetic resonance spectroscopy were used for visualization of PHB granules and chemical confirmation of PHAs in cell extracts, respectively. We report for the first time the production of PHAs by Halococcus sp. (Halococcus morrhuae DSM 1307T, Halococcus saccharolyticus DSM 5350T, Halococcus salifodinae DSM 8989T, Halococcus dombrowskii DSM 14522T, Halococcus hamelinensis JCM 12892T, Halococcus qingdaonensis JCM 13587T), Halorubrum sp. (Hrr. coriense DSM 10284T, Halorubrum chaoviator DSM 19316T, Hrr. chaoviator strains NaxosII and AUS-1), haloalkaliphiles (Natronobacterium gregoryi NCMB 2189T, Natronococcus occultus DSM 3396T) and Halobacterium noricense DSM 9758T. No PHB was detected in Halobacterium salinarum NRC-1 ATCC 700922, Hbt. salinarum R1 and Haloferax volcanii DSM 3757T. Most species synthesized PHAs when growing in synthetic as well as in complex medium. The polyesters were generally composed of PHB and poly-ß-hydroxybutyrate-co-3-hydroxyvalerate (PHBV). Available genomic data suggest the absence of PHA synthesis in some haloarchaea and in all other Euryarchaeota and Crenarchaeota. Homologies between haloarchaeal and bacterial PHA synthesizing enzymes had indicated to some authors probable horizontal gene transfer, which, considering the data obtained in this study, may have occurred already before Permian times. Electronic supplementary material The online version of this article (doi:10.1007/s00253-010-2611-6) contains supplementary material, which is available to authorized users.
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              Integrated analysis of gene expression and metabolic fluxes in PHA-producing Pseudomonas putida grown on glycerol

              Background Given its high surplus and low cost, glycerol has emerged as interesting carbon substrate for the synthesis of value-added chemicals. The soil bacterium Pseudomonas putida KT2440 can use glycerol to synthesize medium-chain-length poly(3-hydroxyalkanoates) (mcl-PHA), a class of biopolymers of industrial interest. Here, glycerol metabolism in P. putida KT2440 was studied on the level of gene expression (transcriptome) and metabolic fluxes (fluxome), using precisely adjusted chemostat cultures, growth kinetics and stoichiometry, to gain a systematic understanding of the underlying metabolic and regulatory network. Results Glycerol-grown P. putida KT2440 has a maintenance energy requirement [0.039 (mmolglycerol (gCDW h)−1)] that is about sixteen times lower than that of other bacteria, such as Escherichia coli, which provides a great advantage to use this substrate commercially. The shift from carbon (glycerol) to nitrogen (ammonium) limitation drives the modulation of specific genes involved in glycerol metabolism, transport electron chain, sensors to assess the energy level of the cell, and PHA synthesis, as well as changes in flux distribution to increase the precursor availability for PHA synthesis (Entner–Doudoroff pathway and pyruvate metabolism) and to reduce respiration (glyoxylate shunt). Under PHA-producing conditions (N-limitation), a higher PHA yield was achieved at low dilution rate (29.7 wt% of CDW) as compared to a high rate (12.8 wt% of CDW). By-product formation (succinate, malate) was specifically modulated under these regimes. On top of experimental data, elementary flux mode analysis revealed the metabolic potential of P. putida KT2440 to synthesize PHA and identified metabolic engineering targets towards improved production performance on glycerol. Conclusion This study revealed the complex interplay of gene expression levels and metabolic fluxes under PHA- and non-PHA producing conditions using the attractive raw material glycerol as carbon substrate. This knowledge will form the basis for the development of future metabolically engineered hyper-PHA-producing strains derived from the versatile bacterium P. putida KT2440.
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                Author and article information

                Contributors
                Role: ND
                Role: ND
                Role: ND
                Role: ND
                Journal
                agro
                Scientia Agropecuaria
                Scientia Agropecuaria
                Universidad Nacional de Trujillo. Facultad de Ciencias Agropecuarias (Trujillo, , Peru )
                2077-9917
                April 2017
                : 8
                : 2
                : 109-118
                Affiliations
                [02] Chulucanas orgnameUniversidad Católica Sedes Sapientiae orgdiv1Fac. de Ingeniería Agraria Peru
                [01] Lambayeque orgnameUniversidad Nacional Pedro Ruiz Gallo orgdiv1Fac. de Ciencias Biológicas Peru
                Article
                S2077-99172017000200003
                10.17268/sci.agropecu.2017.02.03
                9b85ca87-f2e4-40c4-8a9d-257de76c5212

                This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

                History
                : 13 January 2017
                : 16 June 2017
                Page count
                Figures: 0, Tables: 0, Equations: 0, References: 45, Pages: 10
                Product

                SciELO Peru


                bioplásticos,Halophilic bacteria,PHA,bioplastics,biopolymers,starch,Bacterias halófilas,biopolímeros,almidón

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