María del Carmen Guerrero

Biochemical diversity of carboxyl Esterases and Lipases from Lake Arreo (Spain) : a metagenomic approach

ABSTRACT The esterases and lipases from the α/β hydrolase superfamily exhibit an enormous sequence diversity, fold plasticity, and activities. Here, we present the comprehensive sequence and biochemical analyses of seven distinct esterases and lipases from the metagenome of Lake Arreo, an evaporite karstic lake in Spain (42�46′N, 2�59′W; altitude, 655 m). Together with oligonucleotide usage patterns and BLASTP analysis, our study of esterases/lipases mined from Lake Arreo suggests that its sediment contains moderately halophilic and cold-adapted proteobacteria containing DNA fragments of distantly related plasmids or chromosomal genomic islands of plasmid and phage origins. This metagenome encodes esterases/lipases with broad substrate profiles (tested over a set of 101 structurally diverse esters) and habitat-specific characteristics, as they exhibit maximal activity at alkaline pH (8.0 to 8.5) and temperature of 16 to 40�C, and they are stimulated (1.5 to 2.2 times) by chloride ions (0.1 to 1.2 M), reflecting an adaptation to environmental conditions. Our work provides further insights into the potential significance of the Lake Arreo esterases/lipases for biotechnology processes (i.e., production of enantiomers and sugar esters), because these enzymes are salt tolerant and are active at low temperatures and against a broad range of substrates. As an example, the ability of a single protein to hydrolyze triacylglycerols, (non)halogenated alkyl and aryl esters, cinnamoyl and carbohydrate esters, lactones, and chiral epoxides to a similar extent was demonstrated.

ECOSYSTEM METABOLISM IN A MEDITERRANEAN SHALLOW LAKE (LAGUNA DE SANTA OLALLA, DOÑANA NATIONAL PARK, SW SPAIN)

Santa Olalla is a Mediterranean permanent coastal shallow lake, with high annual solar irradiation and warm water temperature throughout the year. Ecosystem metabolism, chlorophyll a concentrations, and physical and chemical features were studied in the Santa Olalla lake from March 1998 to February 2000. Gross primary productivity (GPP) and community respiration (CR) were determined using a modification of the one-station diel oxygen change method. Chlorophyll a, Secchi depth, and total phosphorous and nitrogen reveal Santa Olalla to be a hypereutrophic system. Values of GPP and CR were very high (average 7.88 g O2 m−2 d−1 and 8.52 g O2 m−2 d−1, respectively). Principal components analysis and a multiple regression model showed photosynthetically active radiation and soluble reactive phosphorus to be the main factors that control primary production in Santa Olalla. Also, the annual and interannual variation of water level in Santa Olalla seems to be an important factor that influences the ecological processes of this system. Mean net daily metabolism during a hydrologic period was not statistically different from 0 (0.008 g O2 m−2 d−1, p=0.99), and the average P/R ratio was not statistically different from 1 (1.164, p=0.343). These results suggest that the primary productivity was balanced with community respiration in Santa Olalla lake. The high values of metabolic rates and low nutrient concentrations in this aquatic ecosystem suggest a quick recycling of nutrients, which were assimilated almost instantaneously after mineralization.

DNA map of mutations at the scute locus of Drosophila melanogaster

The achaete‐scute gene complex (AS‐C) of Drosophila melanogaster is involved in the differentiation of innervated elements in the adult (chaetes) and in the embryo (central nervous system). Genetically, the AS‐C is subdivided into four regions: achaete, scute α, lethal of scute, and scute β. Using a previously cloned fragment of scute DNA, we have now cloned 62 kb of wild‐type DNA from the scute region. No repetitive sequences have been detected in this stretch of DNA. Of 16 scute mutants with chromosomal rearrangements studied (inversions, deletions, and translocations), nine, included genetically in scute β, have breakpoints in the cloned region. The remaining rearrangements, which genetically correspond to scute α, map outside and to the left of the cloned region. Of nine scute ‘point mutants’ studied, eight have large DNA alterations within the cloned region. These alterations include insertions (five) and deletions (three). The DNA alterations found in both ‘point mutants’ and rearrangements are interspersed and scattered over 40 kb. The relationship between the sites of the DNA alterations and the mutant phenotypes are discussed.

Spatiotemporal distribution of microbial communities in a coastal, sandy aquifer system (Doñana, SW Spain).

The aquifer system of Doñana (SW Spain) represents the most important freshwater source in the Doñana Natural Area. Its spatiotemporal dynamics favours the hydrological connection between surface and subsurface ecosystems, and promotes matter fluxes among the different terrestrial and aquatic systems present here. This aquifer has been intensively studied from a hydrogeological point of view but little is known from an ecological perspective. In order to understand the ecological roles played by microbial communities in this system, we conducted a long-term seasonal study of bacterial abundance, cell biomass, bacterial biomass and functional activities over a 2-year period. Bacterial abundance ranged between 2.11 +/- 1.79 x 10(5) and 8.58 +/- 6.99 x 10(7) bacteria mL(-1) groundwater, average cell biomass was estimated to be 77.01 +/- 31.56 fgC and bacterial biomass varied between 8.99 +/- 4.10 x 10(-2) and 5.65 +/- 0.70 microgC mL(-1). Iron-related bacteria showed the highest activities among the functional groups studied. Moreover, among the variables that usually control spatial distributions of microbial communities in aquifer systems, depth did not have a relevant effect on this aquifer, at least in the range of depths studied, but grain size, probably due to its direct effects on hydrogeological parameters, such as permeability or porosity, appeared to exert moderate control, principally in terms of bacterial abundance. Finally, significant seasonal differences in the means of these microbiological variables were also observed; temperature seems to be the main factor controlling the temporal distribution of microbial communities in this aquifer system.

Microbial Activities in a Coastal, Sandy Aquifer System (Doñana Natural Protected Area, SW Spain)

We conducted a seasonal research of the activities of microbial communities in a coastal, sandy aquifer system located in the Doñana Natural Protected Area (SW, Spain). Groundwaters from 30 piezometers were sampled over a two-year period. The proportion of active microbial biomass ranged from 0.02 to 6.36% of the total microbial biomass, while the active microbial biomass ranged from 7.42 10 −3 2.20 10 −4 to 17.30 3.71 ngC mL −1 . Bacterial carbon production, measured through the incorporation of [ 3 H]leucine into cellular proteins, showed a mean value of 0.18 0.72 ngC mL −1 h −1 in all wells and all seasons. Bacterial growth rates ranged from 0.03 to 87.26 days. These activities exhibited spatiotemporal patterns. Temperature and the presence of nutrients and organic matter appear to be important factors controlling these patterns. However, hydrogeological flows, both local and regional, seemed to constitute the most important factor determining these spatiotemporal patterns, probably because the distribution of nutrients in aquifer systems is mainly controlled by these hydrogeological flows. The well-known hydrological flows connecting surface waters and groundwaters in Doñana support the assumption that both water compartments form a unique entity (called hydroecosystem), which functions as a whole. Consequently, not only microbial processes in surface waters can influence ecological processes in groundwaters, the characteristics of surface waters can also be affected by groundwater chemical processes, among others, mediated by the activities of microbial communities.