The harvesting of algae rich in triacylglycerol from an open raceway pond with volume-changes synchronized to the circadian cycle and under limited nitrogen supply is researched. A reduced metabolic model of its non-balanced growth is embedded into a macroscopic balance for simulating growth in such a type of bioreactor. Afterwards, the model is used jointly with optimal control to derive cyclic inlet and outlet flow policies as well as optimal nitrogen levels for different time-horizons and initial physiological states in a case study which uses bibliographic data for the Tisochrysis lutea alga.

Microbiota structure changes throughout the treatment of wastewater according to the prevailing physicochemical characteristics. Under optimal conditions, prokaryotic and eukaryotic assemblages participate in the coordinated sequential elimination of contaminants. The relevance of the identification and quantification of the prevailing microorganisms in treatment plants is based on the fact that they play key roles as biological indicators of plant performance and wastewater quality. In this chapter, we focus on molecular and phenotypic methods as complementary strategies to understand the microbial composition and metabolic activities in wastewater. To identify and quantify prokaryotic and eukaryotic groups of organisms, very useful are molecular methods such as sequencing and phylogenetic analysis with molecular markers (i.e., 16 or 18S rRNA genes), fluorescent in situ hybridization, and metagenomics. Phenotypic methods complement the data obtained by the mentioned techniques. Among them, are the micromorphological analysis and grouping the identified organisms into prokaryotic metabolic groups. In this work, examples of the application of these techniques in urban solid waste leachates are discussed. Additionally, we propose out both approaches as useful tools for providing information for the bioprospecting of microorganisms with potential for biotechnological applications such as bioremediation and production of valuable commercial products, for example, bioplastics and biofuels.

The toxic effects of silver nanoparticles (AgNPs) on the physiology and morphology of the green microalga Chlorella vulgaris were studied. AgNPs were characterized by particle size distribution, zeta potential measurement, and atomic force microscopy (AFM). Chlorella vulgaris was exposed to 90-1440 mu g/L of AgNPs range in Bold's Basal Medium for 96 h. The inhibition of algae growth rate and changes in the concentrations of chlorophyll-a, chlorophyll-b, pheophytin, and carotenoids was determined at the beginning and end of the trial. Cell diameter and volume, carbohydrate, total lipids, and protein content were also determined. Our data strongly suggest that the toxic effects of the AgNPs resulted in concentration and time-dependent. AgNPs altered C. vulgaris growth kinetics and cell metabolism expressed in photosynthetic pigments and biochemical composition. Our study confirmed the cytotoxicity of AgNPs through the algal growth inhibition with an EC50 value of 110 mu g/L. Also, changes of chlorophyll-a, chlorophyll-b, pheophytin, and carotenoids concentrations were observed associated with a color shift from green to pale brown of algae cultures exposed to AgNPs for 96 h. Furthermore, algae cell concentration, diameter, and volume, plus total lipid, protein, and carbohydrates contents in the presence of AgNPs, were significantly altered compared to untreated cells. In synthesis, this study highlighted AgNPs toxic effects on morphological and physiological traits of C. vulgaris and warns about possible impacts on energy flow and aquatic food web structure, and on the transfer efficiency of energy to higher trophic levels.

For both bioprospecting of oleaginous microalgae and routine analysis of lipid levels in these microorganisms, reliable techniques are needed. Methods usually used for these purposes include fluorescent staining of lipids and gravimetric determination. In this research, we evaluate the application of a specific Oil Red O staining technique to detect accumulated lipids in the microalgae Chlorella sp., Nitzschia laevis, and Crypthecodinium cohnii. In addition, the viability to evaluate lipid accumulation levels in these microalgae is also analyzed. This methodology offers a less expensive alternative in the study of lipid accumulation levels in oleaginous microalgae since it requires small amounts of cells and low-complexity analytical equipment for its application.

The effects of commercial formulations of Bacillus thuringiensis var. israelensis (Bti) on non-target organisms are still a matter of debate; in amphibians, the risks of Bti are little known. To evaluate the toxicity of a commercial liquid (aqueous suspension, AS) formulation of Bti (Introban®) on Leptodactylus latrans tadpoles, including median lethal concentration (LC50) and no-and lowest–observed-effect concentrations (NOEC and LOEC, respectively), as well as the possible effects of Bti on oxidative responses, erythrocytes genotoxicity, and histology of the intestines. In the laboratory, tadpoles were exposed to nominal concentrations of 0 (control), 2.5, 5, 10, 20 and 40 mg/L of formulated Bti-AS. Glutathione S-transferase (GST) and catalase (CAT) activities, as well as formation of erythrocyte nuclear abnormalities (ENAs), and histological effect were measured in tadpoles displaying survival rates >85%. L. latrans tadpoles were sensitive to exposure to Bti-AS, reaching 100% mortality after 48 h of exposure at the highest concentration. Bti-AS induced GST and CAT enzymes and genotoxicity (erythrocyte's nuclear abnormalities), and caused intestine's histopathology. Our results demonstrate that toxicity of Bti-AS is dose-dependent for L. latrans tadpoles and that sublethal exposure alters enzymes of oxidative stress, induces genotoxicity, and causes intestine damage. Further research is needed to evaluate the ecotoxicological risk of the massive use of Bti formulations on amphibian populations that commonly used suburban wastewater or urban waterbodies to reproduce and where this biopesticide is frequently applied.

In Trypanosoma cruzi, the modification of thiols by glutathionylation-deglutathionylation and its potential relation to protective, regulatory or signaling functions have been scarcely explored. Herein we characterize a dithiolic glutaredoxin (TcrGrx), a redox protein with deglutathionylating activity, having potential functionality to control intracellular homeostasis of protein and non-protein thiols. The catalytic mechanism followed by TcrGrx was found dependent on thiol concentration. Results suggest that TcrGrx operates as a dithiolic or a monothiolic Grx, depending on GSH concentration. TcrGrx functionality to mediate reduction of protein and non-protein disulfides was studied. TcrGrx showed a preference for glutathionylated substrates respect to protein disulfides. From in vivo assays involving TcrGrx overexpressing parasites, we observed the contribution of the protein to increase the general resistance against oxidative damage and intracellular replication of the amastigote stage. Also, studies performed with epimastigotes overexpressing TcrGrx strongly suggest the involvement of the protein in a cellular pathway connecting an apoptotic stimulus and apoptotic-like cell death. Novel information is presented about the participation of this glutaredoxin not only in redox metabolism but also in redox signaling pathways in T. cruzi. The influence of TcrGrx in several parasite physiological processes suggests novel insights about the protein involvement in redox signaling.

Tryparedoxins (TXNs) are multipurpose oxidoreductases from trypanosomatids that transfer reducing equivalents from trypanothione to various thiol proteins. In Trypanosoma cruzi, two genes coding for TXN-like proteins have been identified: TXNI, previously characterized as a cytoplasmic protein, and TXNII, a putative tail-anchored membrane protein. In this work, we performed a comparative functional characterization of T. cruzi TXNs. Particularly, we cloned the gene region coding for the soluble version of TXNII for its heterologous expression. The truncated recombinant protein (without its 22 C-terminal transmembrane amino acids) showed TXN activity. It was also able to transfer reducing equivalents from trypanothione, glutathione, or dihydrolipoamide to various acceptors, including methionine sulfoxide reductases and peroxiredoxins. The results support the occurrence and functionality of a second tryparedoxin, which appears as a new component in the redox scenario for T. cruzi.

Glutaredoxins play an important role in cellular functionality. A putative dithiol glutaredoxin is encoded in the genome of Trypanosoma cruzi. We cloned the gene and obtained the recombinant protein, which behaved as a typical thioltransferase. Activity was variable and dependent on the nature of reducer or oxidant agent used, or both. Epimastigote extracts exhibited similar activity, suggesting the occurrence of the protein in the parasite. Results support a redox scenario in T. cruzi, with glutaredoxin being involved mainly in reduction of glutathione disulfide as well as in deglutathionylation of target proteins.

Glutathione reductase (E.C.1.8.1.7) was purified from Phaeodactylum tricornutum cells grown axenically in an autotrophic medium. The overall procedure started with preparation of the cell extract and addition of ammonium sulfate to 20% saturation, followed by anion exchange and affinity interaction chromatography(Blue-A- and 2',5'-ADP-Sepharose). Complete purification required native polyacrylamide gel electrophoresisas the final step. The enzyme was purified to homogeneity and functionally characterized. Its native molecular mass was estimated to be 118 kDa; which corresponds to a dimer. The enzyme exhibited a specific activity of 190 U mg(-1) with an optimal activity at pH 8.0 and 32 degrees C. We determined K-m values of 14 mu M and 60 mu M for NADPH and oxidized glutathione, respectively. Products inhibited the enzyme according to a hybrid ping-pong reaction mechanism. After MALDI-TOF analysis, the purified enzyme was unambiguously identified as one of the two proteins annotated as glutathione reductases in the genome of the diatom. The properties of the enzyme help to understand redox metabolic scenarios in P. tricornutum. (C) 2009 Elsevier GmbH. All rights reserved.