Effects of nitrogen sources on the nitrate assimilation in Haloferax mediterranei: growth kinetics and transcriptomic analysis

The haloarchaeon Haloferax mediterranei is able to grow in a defined culture media not only in the presence of inorganic nitrogen salt but also with amino acid as the sole nitrogen source. Assimilatory nitrate and nitrite reductases, respectively, catalyze the first and second reactions. The genes involved in this process are nasA, which encodes nitrate reductase and is found within the operon nasABC, and nasD, which encodes nitrite reductase. These genes are subjected to transcriptional regulation, being repressed in the presence of ammonium and induced with either nitrate or nitrite. This type of regulation has also been described when the amino acids are used as nitrogen source in the minimal media. Furthermore, it has been observed that the microorganism growth depends on nitrogen source, obtaining the lowest growth rate in the presence of nitrate and aspartate. In this paper, we present the results of a comparative study of microorganism growth and transcriptomic analysis of the operon nasABC and gene nasD in different nitrogen sources. The results are the first ever produced in relation to amino acids as nitrogen sources within the Halobacteriaceae family.

Characterisation of chlorate reduction in the haloarchaeon Haloferax mediterranei

Background: Haloferax mediterranei is a denitrifying haloarchaeon using nitrate as a respiratory electron acceptor under anaerobic conditions in a reaction catalysed by pNarGH. Other ions such as bromate, perchlorate and chlorate can also be reduced. Methods: Hfx. mediterranei cells were grown anaerobically with nitrate as electron acceptor and chlorate reductase activity measured in whole cells and purified nitrate reductase. Results: No genes encoding (per)chlorate reductases have been detected either in the Hfx. mediterranei genome or in other haloarchaea. However, a gene encoding a chlorite dismutase that is predicted to be exported across the cytoplasmic membrane has been identified in Hfx. mediterranei genome. Cells did not grow anaerobically in presence of chlorate as the unique electron acceptor. However, cells anaerobically grown with nitrate and then transferred to chlorate-containing growth medium can grow a few generations. Chlorate reduction by the whole cells, as well as by pure pNarGH, has been characterised. No clear chlorite dismutase activity could be detected. Conclusions: Hfx. mediterranei pNarGH has its active site on the outer-face of the cytoplasmic membrane and reacts with chlorate and perchlorate. Biochemical characterisation of this enzymatic activity suggests that Hfx. mediterranei or its pure pNarGH could be of great interest for waste water treatments or to better understand biological chlorate reduction in early Earth or Martian environments. General significance: Some archaea species reduce (per)chlorate. However, results here presented as well as those recently reported by Liebensteiner and co-workers [1] suggest that complete perchlorate reduction in archaea follows different rules in terms of biological reactions.

Denitrification Enzymes-Based Biosensors: the Case of Haloarchaeal Enzymes

In the last two decades, the increase in the use of artificial fertilizers and the disposal of industrial wastes have been the main factors responsible for the progressive increase in nitrate and nitrite levels in groundwater and soil. A variety of analytical strategies have been developed for nitrate and nitrite detection but electrochemical biosensors, which are simple, cheap, easily miniaturized and suitability for real-time detection, are proved to be a powerful tool. Various types of biosensors based on the use of whole cells or on the immobilization of denitrification enzymes have been developed, but their use is limited in environmental analysis under extreme conditions such as brines, acidic or basic wastewaters, salted soils, etc. Extremophilic denitrifying microorganism are good candidates for the development of new nitrate and nitrite biosensors and, in particular, haloarchaeal based biosensors would have advantages over bacterial based biosensors since the microorganisms and the purified denitrifying enzymes tolerate a wide range of temperature and salinity. This work summarizes new highlights on the potential uses of denitrifying haloarchaeal enzymes to make enzyme-based biosensors.

Nitrate reduction at Pt(1 0 0) single crystals and preferentially oriented nanoparticles in neutral media

The electroreduction of nitrate on Pt(1 0 0) electrodes in phosphate buffer neutral solution, pH 7.2, is reported. The sensitivity of the reaction to the crystallographic order of the surface is studied through the controlled introduction of defects by using stepped surfaces with (1 0 0) terraces of different length separated by monoatomic steps, either with (1 1 1) or (1 1 0) symmetry. The results of this study show that nitrate reduction occurs mainly on the well defined (1 0 0) terraces in the potential region where H adsorption starts to decrease, allowing the nitrate anion to access the surface. Adsorbed NO has been detected as a stable intermediate in this media. An oxidation process observed at 0.8 V has been identified as leading to the formation of adsorbed NO and being responsible for a secondary reduction process observed in the subsequent negative scan. Using in situ FTIRS, ammonium was found to be the main product of nitrate reduction. This species can be oxidized at high potentials resulting in adsorbed NO and nitrate (probably with nitrite as intermediate).

Green synthesis of polypyrrole-supported metal catalysts: application to nitrate removal in water

Pt and Pt/Sn catalysts supported on polypyrrole (PPy) have been prepared using Ar plasma to reduce the metal precursors dispersed on the polymer. The PPy support was synthesized by chemical polymerization of pyrrole with FeCl3·6H2O, this leading to the conducting form of the polymer (conductimetric measurements). The Ar plasma treatment produced a partial reduction of platinum ions, anchored as platinum chloro-complexes to the PPy chain, into metallic platinum. A homogeneous distribution of Pt and Sn nanoparticles was observed by TEM. Activity of the PPy-supported catalysts was evaluated in the reduction of aqueous nitrate with H2 at room temperature. Nitrate concentration in water below the maximum acceptable level of 50 mg L−1 was achieved with all catalysts. However, considering not only efficiency in nitrate reduction, but also minimized concentrations of undesired nitrite and ammonium, the monometallic Pt catalyst seems to be the most promising one.