Liliana Beatriz Villegas

Responses of Candida fukuyamaensis RCL-3 and Rhodotorula mucilaginosa RCL-11 to copper stress.

The effect of high Cu(II) concentrations on superoxide dismutase (SOD) and catalase (CAT) activity in Candida fukuyamaensis RCL‐3 and Rhodotorula mucilaginosa RCL‐11, previously isolated from a copper filter at a mine plant in Argentina, was studied. Addition of 0.1, 0.2 and 0.5 mM Cu(II) to the culture medium increased total SOD and CAT activity in both strains. Native polyacrylamide gel electrophoresis revealed two bands with SOD activity for C. fukuyamaensis RCL‐3 and only one for R. mucilaginosa RCL‐11; the three bands corresponded to MnSOD.

Production and partial characterization of bioemulsifier from a chromium-resistant actinobacteria

Surface-active compounds such as synthetic emulsifiers have been used for several decades, both for the degradation of hydrocarbons and increasing desorption of soil-bound metals. However, due to their high toxicity, low degradability, and production costs unaffordable for use in larger ecosystems, synthetic emulsifiers have been gradually replaced by those derived from natural sources such as plants or microbes. In previous studies, the bacterium Streptomyces sp. MC1 has shown the ability to reduce and/or accumulate Cr(VI), a highly promising advance in the development of methods for environmental clean-up of sites contaminated with chromium. Here, new studies on the production of emulsifier from this strain are presented. The cultivation factors that have a significant influence on emulsifier biosynthesis, as well as the interactions among them, were studied by factorial design. Based upon optimization studies, maximum bioemulsifier production was detected in the culture medium having an initial pH of 8 with phosphate 2.0 g L(-1) and Ca(+2) 1.0 g L(-1) added, with an emulsification index about 3.5 times greater compared to the basal value. Interestingly, in the presence of 5.0 g L(-1) Cr(VI), Streptomyces sp. MC1 retained about 65% of its emulsifier production ability. Partially purified emulsifier presented high thermo-stability and partial water solubility. These findings could have promising future prospects for the remediation of organic- and metal-contaminated sites.

Production of bioemulsifiers by Amycolatopsis tucumanensis DSM 45259 and their potential application in remediation technologies for soils contaminated with hexavalent chromium.

In recent years, increasing interest has been shown in the use of bioemulsifiers as washing agents that can enhance desorption of soil-bound metals. However, high production costs derived from the use of expensive substrates for formulation of the fermentation media represent the main challenge for full, large-scale implementation of bioemulsifiers. This work reports on a first study of bioemulsifier production by the actinobacterium Amycolatopsis tucumanensis DSM 45259 using different carbon and nitrogen sources. Preliminary results on the potential use of these compounds as washing agents for soils contaminated with Cu(II) and Cr(VI) are also presented. The best specific production was detected using glycerol and urea as carbon and nitrogen substrates, respectively. However, with all of the substrates used during the batch assay, the bioemulsifiers showed high levels of stability at extreme conditions of pH, temperature, and salt concentration. Under the current assay conditions, the bioemulsifiers were not effective in removing Cu(II) from soil. However, they were able to mediate Cr(VI) recovery, with the removal percentage doubled compared to that seen when using deionized water. These findings appear promising for the development of remediation technologies for hexavalent chromium compounds based upon direct use of these microbial emulsifiers.

Raw sugarcane bagasse as carbon source for xylanase production by Paenibacillus species: a potential degrader of agricultural wastes

Paenibacillus species isolated from a variety of natural sources have shown to be important glycoside hydrolases producers. These enzymes play a key role in bio-refining applications, as they are central biocatalysts for the processing of different types of polymers from vegetal biomass. Xylanase production by three native isolates belonging to the genus Paenibacillus was approached by utilizing mineral-based medium and agricultural by-products as a convenient source to produce biocatalysts suitable for their degradation. While varieties of alkali pretreated sugarcane bagasse were useful substrates for the strains from Paenibacillus genus evaluated, raw sugarcane bagasse was the most effective substrate for endoxylanase production by Paenibacillus sp. AR247. This strain was then selected to further improvement of its enzyme production by means of a two-step statistical approach. It was determined that the carbon source, provided as an inexpensive agro-waste, as well as phosphate and magnesium were the culture media components that most influenced the enzyme production, which was improved three times compared to the screening results.

ICPMS analysis of proteins separated by Native-PAGE: Evaluation of metaloprotein profiles in human synovial fluid with acute and chronic arthritis.

The role of trace elements bound to proteins in the etiology and pathogenesis of rheumatoid arthritis (RA) remains unclear. In this sense, the identification and detection of metalloproteins has a strong and growing interest. Metalloprotein studies are currently carried out by polyacrylamide gel electrophoresis (PAGE) associated to inductively coupled plasma mass spectrometry (ICPMS), and despite that complete information can be obtained for metals such as Fe, Cu and Zn, difficulties due to poor sensitivity for other trace elements such as Sn, As, etc, are currently faced. In the present work, a simple and fast method for the determination of trace metals bound to synovial fluid (SF) proteins was optimized. Proteins from SF (long and short-term RA) were separated in ten fractions by native PAGE, then dissolved in nitric acid and peroxide hydrogen, and analyzed by ICPMS. Fifteen metals were determined in each separated protein fraction (band). Adequate calibration of proteins molecular weight allowed stablishing which protein type were bound to different metals.

Cu(II) removal by Rhodotorula mucilaginosa RCL-11 in sequential batch cultures.

The present study explored the ability of the yeast Rhodotorula mucilaginosa RCL-11 to adapt to increasing Cu(II) concentrations, measuring oxidative stress through superoxide dismutase and catalase activity in two parallel sequential batch assays. One assay was performed in Erlenmeyer flasks without aeration and a second in a fermentor in which the dissolved oxygen was maintained at 30% saturation. Both assays were carried out by increasing Cu(II) concentrations in five sequential steps: 0; 0.1; 0.2; 0.5 and 1 mM. Each assay was incubated at 30 degrees C, 250 rpm and pH 5.5. While growth parameters of R. mucilaginosa RCL-11 decreased 90-95% with increasing Cu(II) concentration in the culture medium, the oxidative stress level increased from 30 to 55% in both assays. Cells grown under controlled oxygen conditions showed 30% more copper bioaccumulation and 10% glucose consumption when compared with cells grown without aeration. SOD activity was higher under controlled than without aeration, whereas CAT activity was similar under both test conditions. Cu(II) bioaccumulation by R. mucilaginosa RCL-11 and a possible increase in this capacity by adaptation of the strain under controlled aeration represent a potential valuable tool for treatment of effluents or water bioremediation with high copper contents.

Prokaryotic and eukaryotic community structure affected by the presence of an acid mine drainage from an abandoned gold mine

The acid mine drainage that originates in the abandoned gold mine in San Luis, Argentina, is released into La Carolina stream. The aim of this study was to determine the influence of this mine drainage on the physicochemical parameters of the area studied and on both prokaryotic and eukaryotic community structure. In addition, specific relationships between microbial taxonomic groups and physicochemical parameters were established. The drainage that flows into La Carolina stream acidifies the stream and increases its sulfate, Zn, Cd and Te concentrations. Microbial analysis showed that prokaryotic community structure is mainly affected by pH values. Actinobacteria and Gammaproteobacteria were abundant in samples characterized by low pH values, while Nitrospirae, Chloroflexi, Deltaproteobacteria, Thaumarchaeota and Euryarchaeota were associated with high concentrations of heavy metals. Otherwise, Alphaproteobacteria was present in samples taken in sunlit areas. Regarding eukaryotic community structure, the sunlight had the greatest impact. Inside the mine, in the absence of light, fungi and protists members were the most abundant microorganisms, while those samples taken in the presence of light displayed algae (green algae and diatoms) as the most abundant ones. After receiving the mine drainage, the stream showed a decrease in the diatom abundance and green algae predominated.

Simultaneous chromate and sulfate removal by Streptomyces sp. MC1. Changes in intracellular protein profile induced by Cr(VI)

The purpose of this study was to investigate the influence of increasing sulfate concentrations on chromium removal, to evaluate the effect of the presence of Cr(VI) on sulfate removal by Streptomyces sp. MC1 and to analyze the differential protein expression profile in the presence of this metal for the identification of proteins repressed or overexpressed. In the presence of Cr(VI) but in the absence of sulfate ions, bacterial growth was negligible, showing the Cr(VI) toxicity for this bacterium. However, the sulfate presence stimulated bacterium growth and Cr(VI) removal, regardless of its concentrations. Streptomyces sp. MC1 showed ability to remove chromium and sulfate simultaneously. Also, the sulfate presence favored the decrease of total chromium concentration from supernatants reaching a decrease of 50% at 48 h. In presence of chromium, seven proteins were down‐expressed and showed homology to proteins involved in protein biosynthesis, energy production and free radicals detoxification while two proteins involved in oxidation‐reduction processes identified as dihydrolipoamide dehydrogenase and S‐adenosyl‐l‐methionine synthase were overexpressed.

Microbial Consortia, a Viable Alternative for Cleanup of Contaminated Soils

The growth population and anthropogenic activity are constantly threatening the environment caused by the accumulation of different kinds of pollutants in the biosphere, especially in soils and sediments. Co-contaminated of environment with toxic organic and inorganic substance is often actually. For the remediation of soils contaminated with mainly petroleum, pesticide and heavy metals, several physical or chemical techniques have been developed inadequately. Inside the technologies “eco-friendly remediation,” the bioremediation have emerged as an option using natural biological activity. Bioremediation are methods where microorganisms degrade one or various pollutants to nontoxic compounds, so working individually or coordinately inside a microbial consortium. A microbial consortium is the natural association of two or more microbial populations of different species, which act together in a complex system. The success of a bioremediation process with pure cultures is very low and restricted. Therefore, use of a microbial consortium appears to be more feasible and reliable.