Exposure of Deinococcus radiodurans to both static magnetic fields and gamma radiation: observation of cell recuperation effects.

The extremophilic bacterium Deinococcus radiodurans displays an extraordinary ability to withstand lethal radiation effects, due to its complex mechanisms for both proteome radiation protection and DNA repair. Published results obtained recently at this laboratory show that D. radiodurans submitted to ionizing radiation results in its DNA being shattered into small fragments which, when exposed to a "static electric field' (SEF), greatly decreases cell viability. These findings motivated the performing of D. radiodurans exposed to gamma radiation, yet exposed to a different exogenous physical agent, "static magnetic fields" (SMF). Cells of D. radiodurans [strain D.r. GY 9613 (R1)] in the exponential phase were submitted to 60Co gamma radiation from a gamma cell. Samples were exposed to doses in the interval 0.5-12.5 kGy, while the control samples were kept next to the irradiation setup. Exposures to SMF were carried out with intensities of 0.08 T and 0.8 T delivered by two settings: (a) a device built up at this laboratory with niobium magnets, delivering 0.08 T, and (b) an electromagnet (Walker Scientific) generating static magnetic fields with intensities from 0.1 to 0.8 T. All samples were placed in a bacteriological incubator at 30 °C for 48 h, and after incubation, a counting of colony forming units was performed. Two sets of cell surviving data were measured, each in triplicate, obtained in independent experiments. A remarkable similarity between the two data sets is revealed, underscoring reproducibility within the 5% range. Appraisal of raw data shows that exposure of irradiated cells to SMF substantially increases their viability. Data interpretation strongly suggests that the increase of D. radiodurans cell viability is a sole magnetic physical effect, driven by a stochastic process, improving the efficiency of the rejoining of DNA fragments, thus increasing cell viability. A type of cut-off dose is identified at 10 kGy, above which the irradiated cellular system loses recovery and the cell survival mechanism collapses.

Comparison of mono-rhamnolipids and di-rhamnolipids on microbial enhanced oil recovery (MEOR) applications.

Rhamnolipids (RMLs) have more effectiveness for specific uses according to their homologue proportions. Thus, the novelty of this work was to compare mono-RMLs and di-RMLs physicochemical properties on microbial enhanced oil recovery (MEOR) applications. For this, RML produced by three strains of Pseudomonas aeruginosa containing different homologues proportion were used: a mainly mono-RMLs producer (mono-RMLs); a mainly di-RMLs producer (di-RMLs), and the other one that produces relatively balanced amounts of mono-RML and di-RML homologues (mono/di-RML). For mono-RML, the most abundant molecules were Rha-C10 C10 (m/z 503.3), for di-RML were RhaRha-C10 C10 (m/z 649.4) and for Mono/di-RML were Rha-C10 C10 (m/z 503.3) and RhaRha-C10 C10 (m/z 649.4). All RMLs types presented robustness under high temperature and variation of salinity and pH, and high ability for oil displacement, foam stability, wettability reversal and were classified as safe for environment according to the European Union Directive No. 67/548/EEC. For all these properties, it was observed a highlight for mono-RML. Mono-RML presented the lowest surface tension (26.40 mN/m), interfacial tension (1.14 mN/m), and critical micellar concentration (CMC 27.04 mg/L), the highest emulsification index (EI24 100%) and the best wettability reversal (100% with 25 ppm). In addition, mono-RML showed the best acute toxicity value (454 mg/L), making its application potential even more attractive. Based on the results, it was concluded that all RMLs homologues studied have potential for MEOR applications. However, results showed that mono-RML stood out and have the best mechanism of oil incorporation in micelles due their most effective surface-active physicochemical features.

A non-naturally-occurring P(3HB-co-3HAMCL) is produced by recombinant Pseudomonas sp. from an unrelated carbon source.

Pseudomonas sp. PHA- was used as host for PHA biosynthesis genes from Aeromonas sp. to produce 3HB-co-3HAMCL from glucose with no supply of co-substrates. A non-naturally-occurring PHA composed mainly of 3HB, 3HHx and 3HD (3HO, 3HDdΔ5 and 3HDd monomers were detected in smaller amounts) was obtained. The polymer was extracted using two different solvents (acetone and chloroform) and subject to the following characterization tests: FTIR, DSC, TGA and GPC. The latter suggests a block copolymer since a single and narrow elution peak was observed for each sample. The DSC results ruled out the possibility of a random copolymer and agrees with a single copolymer composed of two blocks: one with the typical composition of PHAMCL produced by Pseudomonas and another containing 3HB and 3HHx with a high 3HHx molar fraction. Thus, this study increases the perspectives of P(3HB-co-3HAMCL) production from carbohydrates as the sole carbon source.

xylA and xylB overexpression as a successful strategy for improving xylose utilization and poly-3-hydroxybutyrate production in Burkholderia sacchari.

Despite the versatility and many advantages of polyhydroxyalkanoates as petroleum-based plastic substitutes, their higher production cost compared to petroleum-based polymers has historically limited their large-scale production. One appealing approach to reducing production costs is to employ less expensive, renewable feedstocks. Xylose, for example is an abundant and inexpensive carbon source derived from hemicellulosic residues abundant in agro-industrial waste (sugarcane bagasse hemicellulosic hydrolysates). In this work, the production of poly-3-hydroxybutyrate P(3HB) from xylose was studied to develop technologies for conversion of agro-industrial waste into high-value chemicals and biopolymers. Specifically, this work elucidates the organization of the xylose assimilation operon of Burkholderia sacchari, a non-model bacterium with high capacity for P(3HB) accumulation. Overexpression of endogenous xylose isomerase and xylulokinase genes was successfully assessed, improving both specific growth rate and P(3HB) production. Compared to control strain (harboring pBBR1MCS-2), xylose utilization in the engineered strain was substantially improved with 25% increase in specific growth rate, 34% increase in P(3HB) production, and the highest P(3HB) yield from xylose reported to date for B. sacchari (YP3HB/Xil = 0.35 g/g). This study highlights that xylA and xylB overexpression is an effective strategy to improve xylose utilization and P(3HB) production in B. sacchari.

Combining molecular and bioprocess techniques to produce poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) with controlled monomer composition by Burkholderia sacchari.

Biopolymers as polyhydroxyalkanoates (PHA) composed by different co-monomers 3-hydroxybutyrate and 3-hydroxyhexanoate [P(3HB-co-3HHx)] has attracted interest since its properties are similar to low density polyethylene. Burkholderia sacchari produces this copolymer with a very low 3HHx molar fraction, about 2 mol%. B. sacchari mutant (unable to produce polymer) was engineered to host PHA biosynthesis genes (phaPCJ) from Aeromonas sp. In addition, a two-step bioprocess to increase biopolymer production was developed. The combination of these techniques resulted in the production of P(3HB-co-3HHx) with 3HHx content up to 20 mol%. The PHA content was about 78% of dry biomass, resulting in PHA volumetric productivities around 0.45gl-1h-1. The P(3HB-co-3HHx) containing 20 mol% of 3HHx presented an elongation at brake of 945%, higher than reported before for this PHA composition. Here we have described an approach to increase 3HHx content into the copolymer, allowing the precise control of the 3HHx molar fractions.

A suitable procedure to choose antimicrobials as controlling agents in fermentations performed by bacteria

This work evaluated the influence of nitrofurantoin, erythromycin and streptomycin at 50, 25 and 12,5 (per cent) of the minimal inhibitory concentration (MIC) on maximum specific growth rate (µmax) and specific polymer accumulation rate (µPHB) of "Alcaligenes eutrophus", considered resistant to those antimicrobials. Nitrofurantoin strongly affected µmax even at 50(per cent) MIC. Streptomycin moderately affected µmax only at 50(per cent) MIC. Nitrofurantoin showed the most harmful effect on µPHB when 50(per cent) MIC was applied and erythromycin was not harmful.