Olga Sánchez

Genome analysis of the proteorhodopsin-containing marine bacterium Polaribacter sp. MED152 (Flavobacteria)

Analysis of marine cyanobacteria and proteobacteria genomes has provided a profound understanding of the life strategies of these organisms and their ecotype differentiation and metabolisms. However, a comparable analysis of the Bacteroidetes, the third major bacterioplankton group, is still lacking. In the present paper, we report on the genome of Polaribacter sp. strain MED152. On the one hand, MED152 contains a substantial number of genes for attachment to surfaces or particles, gliding motility, and polymer degradation. This agrees with the currently assumed life strategy of marine Bacteroidetes. On the other hand, it contains the proteorhodopsin gene, together with a remarkable suite of genes to sense and respond to light, which may provide a survival advantage in the nutrient-poor sun-lit ocean surface when in search of fresh particles to colonize. Furthermore, an increase in CO2 fixation in the light suggests that the limited central metabolism is complemented by anaplerotic inorganic carbon fixation. This is mediated by a unique combination of membrane transporters and carboxylases. This suggests a dual life strategy that, if confirmed experimentally, would be notably different from what is known of the two other main bacterial groups (the autotrophic cyanobacteria and the heterotrophic proteobacteria) in the surface oceans. The Polaribacter genome provides insights into the physiological capabilities of proteorhodopsin-containing bacteria. The genome will serve as a model to study the cellular and molecular processes in bacteria that express proteorhodopsin, their adaptation to the oceanic environment, and their role in carbon-cycling.

Winter‐to‐summer changes in the composition and single‐cell activity of near‐surface Arctic prokaryotes

We collected surface samples in Franklin Bay (Western Arctic) from ice-covered to ice-free conditions, to determine seasonal changes in the identity and in situ activity of the prokaryotic assemblages. Catalysed reported fluorescence in situ hybridization was used to quantify the abundance of different groups, and combined with microautoradiography to determine the fraction of active cells taking up three substrates: glucose, amino acids and ATP. In surface waters, Archaea accounted for 16% of the total cell count in winter, but decreased to almost undetectable levels in summer, when Bacteria made up 97% of the total cell count. Alphaproteobacteria were the most abundant group followed by Bacteroidetes (average of 34% and 14% of total cell counts respectively). Some bacterial groups appearing in low abundances (< 10% of total cell counts), such as Betaproteobacteria, Roseobacter and Gammaproteobacteria, showed a high percentage of active cells. By contrast, more abundant groups, such as SAR11 or Bacteroidetes, had a lower percentage of active cells in the uptake of the substrates tested. Archaea showed low heterotrophic activity throughout the year. In comparison with temperate oceans, the percentage of active Bacteria in the uptake of the substrates was relatively high, even during the winter season.

Comparison of different denaturing gradient gel electrophoresis primer sets for the study of marine bacterioplankton communities.

ABSTRACT An annual seasonal cycle of composition of a bacterioplankton community in an oligotrophic coastal system was studied by denaturing gradient gel electrophoresis (DGGE) using five different primer sets. Analysis of DGGE fingerprints showed that primer set 357fGC-907rM grouped samples according to seasons. Additionally, we used the set of 16S rRNA genes archived in the RDPII database to check the percentage of perfect matches of each primer for the most abundant bacterial groups inhabiting coastal plankton communities. Overall, primer set 357fGC-907rM was the most suitable for the routine use of PCR-DGGE analyses in this environment.

Microbial communities from different types of natural wastewater treatment systems: Vertical and horizontal flow constructed wetlands and biofilters

The prokaryotic microbial communities (Bacteria and Archaea) of three different systems operating in Denmark for the treatment of domestic wastewater (horizontal flow constructed wetlands (HFCW), vertical flow constructed wetlands (VFCW) and biofilters (BF)) was analysed using endpoint PCR followed by Denaturing Gradient Gel Electrophoresis (DGGE). Further sequencing of the most representative bacterial bands revealed that diverse and distinct bacterial communities were found in each system unit, being γ-Proteobacteria and Bacteroidetes present mainly in all of them, while Firmicutes was observed in HFCW and BF. Members of the Actinobacteria group, although found in HFCW and VFCW, seemed to be more abundant in BF units. Finally, some representatives of α, β and δ-Proteobacteria, Acidobacteria and Chloroflexi were also retrieved from some samples. On the other hand, a lower archaeal diversity was found in comparison with the bacterial population. Cluster analysis of the DGGE bacterial band patterns showed that community structure was related to the design of the treatment system and the organic matter load, while no clear relation was established between the microbial assemblage and the wastewater influent.

Molecular characterization of an oil-degrading cyanobacterial consortium

Recent studies have shown that the cyanobacterium Microcoleus chthonoplastes forms a consortium with heterotrophic bacteria present within the cyanobacterial sheath. These studies also show that this consortium is able to grow in the presence of crude oil, degrading aliphatic heterocyclic organo-sulfur compounds as well as alkylated monocyclic and polycyclic aromatic hydrocarbons. In this work, we characterize this oil-degrading consortium through the analysis of the 16S rRNA gene sequences. We performed the study in cultures of Microcoleus grown in mineral medium and in cultures of the cyanobacterium grown in mineral medium supplemented with crude oil. The results indicate that most of the clones found in the polluted culture correspond to well-known oil-degrading and nitrogen-fixing microorganisms, and belong to different phylogenetic groups, such as the Alpha, Beta, and Gamma subclasses of Proteobacteria, and the Cytophaga/Flavobacteria/Bacteroides group. The control is dominated by one predominant organism (88% of the clones) closely affiliated to Pseudoxanthomonas mexicana (similarity of 99.8%). The presence of organisms closely related to well-known nitrogen fixers such as Rhizobium and Agrobacterium suggests that at least some of the cyanobacteria-associated heterotrophic bacteria are responsible for nitrogen fixation and degradation of hydrocarbon compounds inside the polysaccharidic sheath, whereas Microcoleus provides a habitat and a source of oxygen and organic matter.

Transient storage of electrical charge in biofilms of Shewanella oneidensis MR-1 growing in a microbial fuel cell.

Current output of microbial fuel cells (MFCs) depends on a number of engineering variables mainly related to the design of the fuel cell reactor and the materials used. In most cases the engineering of MFCs relies on the premise that for a constant biomass, current output correlates well with the metabolic activity of the cells. In this study we analyze to what extent, MFC output is also affected by the mode of operation, emphasizing how discontinuous operation can affect temporal patterns of current output. The experimental work has been carried out with Shewanella oneidensis MR-1, grown in conventional two-chamber MFCs subject to periodic interruptions of the external circuit. Our results indicate that after closure of the external circuit, current intensity shows a peak that decays back to basal values. The result suggests that the MFC has the ability to store charge during open circuit situations. Further studies using chronoamperometric analyses were carried out using isolated biofilms of Shewanella oneidensis MR-1 developed in a MFC and placed in an electrochemistry chamber in the presence of an electron donor. The results of these studies indicate that the amount of excess current over the basal level released by the biofilm after periods of circuit disconnection is proportional to the duration of the disconnection period up to a maximum of approximately 60 min. The results indicate that biofilms of Shewanella oneidensis MR-1 have the ability to store charge when oxidizing organic substrates in the absence of an external acceptor.

Key design factors affecting microbial community composition and pathogenic organism removal in horizontal subsurface flow constructed wetlands

Constructed wetlands constitute an interesting option for wastewater reuse since high concentrations of contaminants and pathogenic microorganisms can be removed with these natural treatment systems. In this work, the role of key design factors which could affect microbial removal and wetland performance, such as granular media, water depth and season effect was evaluated in a pilot system consisting of eight parallel horizontal subsurface flow (HSSF) constructed wetlands treating urban wastewater from Les Franqueses del Vallès (Barcelona, Spain). Gravel biofilm as well as influent and effluent water samples of these systems were taken in order to detect the presence of bacterial indicators such as total coliforms (TC), Escherichia coli, fecal enterococci (FE), Clostridium perfringens, and other microbial groups such as Pseudomonas and Aeromonas. The overall microbial inactivation ratio ranged between 1.4 and 2.9 log-units for heterotrophic plate counts (HPC), from 1.2 to 2.2 log units for total coliforms (TC) and from 1.4 to 2.3 log units for E. coli. The presence of fine granulometry strongly influenced the removal of all the bacterial groups analyzed. This effect was significant for TC (p=0.009), E. coli (p=0.004), and FE (p=0.012). Shallow HSSF constructed wetlands were more effective for removing Clostridium spores (p=0.039), and were also more efficient for removing TC (p=0.011) and E. coli (p=0.013) when fine granulometry was used. On the other hand, changes in the total bacterial community from gravel biofilm were examined by using denaturing gradient gel electrophoresis (DGGE) and sequencing of polymerase chain reaction (PCR)-amplified fragments of the 16S rRNA gene recovered from DGGE bands. Cluster analysis of the DGGE banding pattern from the different wetlands showed that microbial assemblages separated according to water depth, and sequences of different phylogenetic groups, such as Alpha, Beta and Delta-Proteobacteria, Nitrospirae, Bacteroidetes, Acidobacteria, Firmicutes, Synergistetes and Deferribacteres could be retrieved from DGGE bands.

Optical emission characterization of CH4+H2 discharges for diamond deposition

Methane and hydrogen discharges has been studied at different discharge frequencies (35 kHz, 13.56 MHz, and 2.45 GHz) and feeding gas ratios (up to 100% of methane) during diamond and diamond‐like deposition by plasma chemical vapor deposition techniques. Optical emission spectroscopy shows that the intensity of atomic hydrogen line (Hα) is the highest at the microwave frequency (2.45 GHz). In addition, at this frequency and low methane concentrations (<7.5%) the emission of CH+ species is also detected, which has been associated to the presence of the diamond phase in the films. On the contrary, at the lower frequencies (35 kHz and 13.56 MHz), the emission spectra are dominated by neutral CH species that are supposed to be the precursor species in the diamond‐like films deposited at these frequencies.

Deposition of TiN/AlN bilayers on a rotating substrate by reactive sputtering

Abstract A balanced dual-cathode reactive magnetron sputtering system has been developed, which allows the growth of TiN/AlN bilayers from pure titanium and aluminium targets in a mixture of argon and nitrogen gases. One important property of the TiN/AlN coatings is that they combine two important requirements for protective coatings: good adhesion properties of the TiN films and the chemical stability of the AlN films. The chemical composition of the grown layers has been studied by Auger electron spectroscopy (AES), elastic recoil detection analysis (ERDA) and Rutherford backscattering spectroscopy (RBS) techniques, and nitrogen incorporation in the nitrides has been correlated to the argon/nitrogen ratio in the sputtering gas. Structural properties have been studied by X-ray diffraction (XRD), and the results showed the presence of cubic TiN and hexagonal AlN phases in the grown bilayer. Mechanical properties have been extracted from nano-indentation measurements, giving hardness values for TiN/AlN bilayer in the range of the reported values for TiN coatings.

A new non-aerated illuminated packed-column reactor for the development of sulfide-oxidizing biofilms

Abstract This paper describes an illuminated reactor that allows the spontaneous development of biofilms aimed at the treatment of sulfide-containing streams. The reactor operates as a sulfidostat and is composed of an illuminated packed-column, in which microorganisms are exposed to constant low substrate concentrations, thereby avoiding inhibition due to high sulfide concentrations. The control system allows highly polluted streams to be oxidized by the microbial biofilm while ensuring the quality of the effluent produced. Both monospecies and multispecies biofilms have been developed. Biofilms undergo changes in light irradiance and sulfide load while providing a consistent reduction of the sulfide levels, down to micromolar concentrations. Both types of biofilm developed differ from stirred reactors in that their specific activities are lower, constituting systems with a slow dynamic behavior and, therefore, they are less sensitive to sudden disturbances.