Enhanced tolerance to naphthalene and enhanced rhizoremediation performance for Pseudomonas putida KT2440 via the NAH7 catabolic plasmid.

In this work, we explore the potential use of the Pseudomonas putida KT2440 strain for bioremediation of naphthalene-polluted soils. Pseudomonas putida strain KT2440 thrives in naphthalene-saturated medium, establishing a complex response that activates genes coding for extrusion pumps and cellular damage repair enzymes, as well as genes involved in the oxidative stress response. The transfer of the NAH7 plasmid enables naphthalene degradation by P. putida KT2440 while alleviating the cellular stress brought about by this toxic compound, without affecting key functions necessary for survival and colonization of the rhizosphere. Pseudomonas putida KT2440(NAH7) efficiently expresses the Nah catabolic pathway in vitro and in situ, leading to the complete mineralization of [(14)C]naphthalene, measured as the evolution of (14)CO(2), while the rate of mineralization was at least 2-fold higher in the rhizosphere than in bulk soil.

Effect of electrokinetics on the bioaccessibility of polycyclic aromatic hydrocarbons in polluted soils.

Bioaccessibility is one of the most relevant aspects to be considered in the restoration of soils using biological technologies. Polycyclic aromatic hydrocarbons (PAH) usually have residual fractions that are resistant to biodegradation at the end of the biological treatment. In some situations, these residual concentrations could still be above legal standards. Here, we propose that the available knowledge about electroremediation technologies could be applied to enhance bioremediation of soils polluted with PAH. The main objective of this study was to show that a previous electrokinetic treatment could reduce the PAH residual fractions when the soil is subsequently treated by means of a bioremediation process. The approach involved the electrokinetic treatment of PAH-polluted soils at a potential drop of 0.9 to 1.1 V/cm and the subsequent estimations of bioaccessibility of residual PAHs after slurry-phase biodegradation. Bioaccessibility of PAH in two creosote-polluted soils (clay and loamy sand, total PAH content averaging 300 mg/kg) previously treated with an electric field in the presence of nonionic surfactant Brij 35 was often higher than in untreated controls. For example, total PAH content remaining in clay soil after bioremediation was only 62.65 +/- 4.26 mg/kg, whereas a 7-d electrokinetic pretreatment had, under the same conditions, a residual concentration of 29.24 +/- 1.88 mg/kg after bioremediation. Control treatments without surfactant indicated that the electrokinetic treatment increased bioaccessibility of PAHs. A different manner of electric field implementation (continuous current vs. current reversals) did not induce changes in PAH bioaccessibility. We suggest that this hybrid technology may be useful in certain bioremediation scenarios, such as soils rich in clay and black carbon, which show limited success due to bioavailability restrictions, as well as in highly heterogeneous soils.

Metabolic engineering, new antibiotics and biofilm viscoelasticity.

In the following highlight we refer to a number of new advances in the field of Biotechnology that address issues relating to the synthesis of new antibiotics, new biocatalysts and matrices in biofilms.

Dual 14C/residue analysis method to assess the microbial accessibility of native phenanthrene in environmental samples.

The aim of this work was to develop a method to assess the microbial accessibility of native phenanthrene present in soils and sediments. We developed an accelerated biodegradation assay, characterized by (a) inoculation with a sufficient number of phenanthrene-degrading microorganisms, (b) monitoring of the biodegradation activity through 14C-mineralization measurements, and (c) single-step chemical analysis of the native compound in the residue. The use of 14C-labeling allowed the determination of the time period needed for biodegradation of the bioaccessible fraction of the native chemical. The method was tested with environmental samples having a wide range of phenanthrene concentrations, i.e., from background levels (microg kg(-1)) originating in soil from atmospheric deposition, to acute concentrations (g kg(-1)) corresponding to industrial pollution of soils and sediments. The results showed a wide range of bioaccessibility (15-95% of the initial amount). The method can be used for the assessment of bioaccessibility involved in the management of polycyclic aromatic hydrocarbon (PAH) pollution.

Integrating biodegradation and electroosmosis for the enhanced removal of polycyclic aromatic hydrocarbons from creosote-polluted soils.

This paper presents a hybrid technology of soil remediation based on the integration of biodegradation and electroosmosis. We employed soils with different texture (clay soil and loamy sand) containing a mixture of polycyclic aromatic hydrocarbons (PAH) present in creosote, and inoculation with a representative soil bacterium able to degrade fluorene, phenanthrene, fluoranthene, pyrene, anthracene, and benzo[a]pyrene. Two different modes of treatment were prospected: (i) inducing in soil the simultaneous occurrence of biodegradation and electroosmosis in the presence of a biodegradable surfactant, and (ii) treating the soils sequentially with electrokinetics and bioremediation. Losses of PAH due to simultaneous biodegradation and electroosmosis (induced by a continuous electric field) were significantly higher than in control cells that contained the surfactant but no biological activity or no current. The method was especially successful with loamy sand. For example, benzo[a]pyrene decreased its concentration by 50% after 7 d, whereas 22 and 17% of the compound had disappeared as a result of electrokinetic flushing and bioremediation alone, respectively. The use of periodical changes in polarity and current pulses increased by 16% in the removal of total PAH and in up to 30% of specific compounds, including benzo[a]pyrene. With the aim of reaching lower residual levels through bioremediation, an electrokinetic pretreatment was also evaluated as a way to mobilize the less bioaccessible fraction of PAH. Residual concentrations of total biodegradable PAH, remaining after bioremediation in soil slurries, were twofold lower in electrokinetically pretreated soils than in untreated soils. The results indicate that biodegradation and electroosmosis can be successfully integrated to promote the removal of PAH from soil.

Electrokinetic enhancement of phenanthrene biodegradation in creosote-polluted clay soil.

Given the difficulties caused by low-permeable soils in bioremediation, a new electrokinetic technology is proposed, based on laboratory results with phenanthrene, to afford bioremediation of polycyclic aromatic hydrocarbons (PAH) in clay soils. Microbial activity in a clay soil historically polluted with creosote was promoted using a specially designed electrokinetic cell with a permanent anode-to-cathode flow and controlled pH. The rates of phenanthrene losses during treatment were tenfold higher in soil treated with an electric field than in the control cells without current or microbial activity. Results from experiments with Tenax-assisted desorption and mineralization of 14C-labeled phenanthrene indicated that phenanthrene biodegradation was limited by mass-transfer of the chemical. We suggest that the enhancement effect of the applied electric field on phenanthrene biodegradation resulted from mobilization of the PAH and nutrients dissolved in the soil fluids.

Biosurfactant- and biodegradation-enhanced partitioning of polycyclic aromatic hydrocarbons from nonaqueous-phase liquids.

A study was conducted on the effect of two different biological factors, microbial surfactants and biodegradation, on the kinetics of partitioning of polycyclic aromatic hydrocarbons (PAHs) from nonaqueous-phase liquids (NAPLs). The effect of rhamnolipid biosurfactants on partitioning into the aqueous phase of naphthalene, fluorene, phenanthrene, and pyrene, initially dissolved in di-2-ethylhexyl phthalate (DEHP) or 2,2,4,4,6,8,8-heptamethylnonane (HMN), was determined in multiple-solute experiments. Biosurfactants at a concentration above the CMC enhanced the partitioning rate of fluorene, phenanthrene, and pyrene but were ineffective with naphthalene. Enhancement of partitioning was also observed in the presence of suspended humic acid-clay complexes, which simulated the solids often present in the subsurface. Biosurfactants sorbed to the complexes modified PAH partitioning between the NAPL and these solids, increasing the fraction of solid-phase PAH. Biodegradation-driven partitioning was estimated in mineralization experiments with phenanthrene initially present in HMN and three representative soil bacterial strains, differing in their potential adherence to the NAPL. In the three cases, the rates of mineralization were very similar and significantly higher than the abiotic rate of partitioning. Our study suggests that in NAPL-polluted sites, partitioning of PAH may be efficiently enhanced by in situ treatments involving the use of biosurfactants and biodegradation.