Arsenic Nanoparticles Trigger Apoptosis via Anoikis Induction in OECM-1 Cells.

Arsenic compounds have been used as therapeutic alternatives for several diseases including cancer. In the following work, we obtained arsenic nanoparticles (AsNPs) produced by an anaerobic bacterium from the Salar de Ascotán, in northern Chile, and evaluated their effects on the human oral squamous carcinoma cell line OECM-1. Resazurin reduction assays were carried out on these cells using 1-100 µM of AsNPs, finding a concentration-dependent reduction in cell viability that was not observed for the non-tumoral gastric mucosa-derived cell line GES-1. To establish if these effects were associated with apoptosis induction, markers like Bcl2, Bax, and cleaved caspase 3 were analyzed via Western blot, executor caspases 3/7 via luminometry, and DNA fragmentation was analyzed by TUNEL assay, using 100 µM cisplatin as a positive control. OECM-1 cells treated with AsNPs showed an induction of both extrinsic and intrinsic apoptotic pathways, which can be explained by a significant decrease in P-Akt/Akt and P-ERK/ERK relative protein ratios, and an increase in both PTEN and p53 mRNA levels and Bit-1 relative protein levels. These results suggest a prospective mechanism of action for AsNPs that involves a potential interaction with extracellular matrix (ECM) components that reduces cell attachment and subsequently triggers anoikis, an anchorage-dependent type of apoptosis.

Looking for the mechanism of arsenate respiration of Fusibacter sp. strain 3D3, independent of ArrAB.

The literature has reported the isolation of arsenate-dependent growing microorganisms which lack a canonical homolog for respiratory arsenate reductase, ArrAB. We recently isolated an arsenate-dependent growing bacterium from volcanic arsenic-bearing environments in Northern Chile, Fusibacter sp. strain 3D3 (Fas) and studied the arsenic metabolism in this Gram-positive isolate. Features of Fas deduced from genome analysis and comparative analysis with other arsenate-reducing microorganisms revealed the lack of ArrAB coding genes and the occurrence of two arsC genes encoding for putative cytoplasmic arsenate reductases named ArsC-1 and ArsC-2. Interestingly, ArsC-1 and ArsC-2 belong to the thioredoxin-coupled family (because of the redox-active disulfide protein used as reductant), but they conferred differential arsenate resistance to the E. coli WC3110 ΔarsC strain. PCR experiments confirmed the absence of arrAB genes and results obtained using uncouplers revealed that Fas growth is linked to the proton gradient. In addition, Fas harbors ferredoxin-NAD+ oxidoreductase (Rnf) and electron transfer flavoprotein (etf) coding genes. These are key molecular markers of a recently discovered flavin-based electron bifurcation mechanism involved in energy conservation, mainly in anaerobic metabolisms regulated by the cellular redox state and mostly associated with cytoplasmic enzyme complexes. At least three electron-bifurcating flavoenzyme complexes were evidenced in Fas, some of them shared in conserved genomic regions by other members of the Fusibacter genus. These physiological and genomic findings permit us to hypothesize the existence of an uncharacterized arsenate-dependent growth metabolism regulated by the cellular redox state in the Fusibacter genus.

From Laboratory towards Industrial Operation: Biomarkers for Acidophilic Metabolic Activity in Bioleaching Systems.

In the actual mining scenario, copper bioleaching, mainly raw mined material known as run-of-mine (ROM) copper bioleaching, is the best alternative for the treatment of marginal resources that are not currently considered part of the profitable reserves because of the cost associated with leading technologies in copper extraction. It is foreseen that bioleaching will play a complementary role in either concentration-as it does in Minera Escondida Ltd. (MEL)-or chloride main leaching plants. In that way, it will be possible to maximize mines with installed solvent-extraction and electrowinning capacities that have not been operative since the depletion of their oxide ores. One of the main obstacles for widening bioleaching technology applications is the lack of knowledge about the key events and the attributes of the technology's critical events at the industrial level and mainly in ROM copper bioleaching industrial operations. It is relevant to assess the bed environment where the bacteria-mineral interaction occurs to learn about the limiting factors determining the leaching rate. Thus, due to inability to accurately determine in-situ key variables, their indirect assessment was evaluated by quantifying microbial metabolic-associated responses. Several candidate marker genes were selected to represent the predominant components of the microbial community inhabiting the industrial heap and the metabolisms involved in microbial responses to changes in the heap environment that affect the process performance. The microbial community's predominant components were Acidithiobacillus ferrooxidans, At. thiooxidans, Leptospirillum ferriphilum, and Sulfobacillus sp. Oxygen reduction, CO2 and N2 fixation/uptake, iron and sulfur oxidation, and response to osmotic stress were the metabolisms selected regarding research results previously reported in the system. After that, qPCR primers for each candidate gene were designed and validated. The expression profile of the selected genes vs. environmental key variables in pure cultures, column-leaching tests, and the industrial bioleaching heap was defined. We presented the results obtained from the industrial validation of the marker genes selected for assessing CO2 and N2 availability, osmotic stress response, as well as ferrous iron and sulfur oxidation activity in the bioleaching heap process of MEL. We demonstrated that molecular markers are useful for assessing limiting factors like nutrients and air supply, and the impact of the quality of recycled solutions. We also learned about the attributes of variables like CO2, ammonium, and sulfate levels that affect the industrial ROM-scale operation.

An indigenous bacterium with enhanced performance of microbially-induced Ca-carbonate biomineralization under extreme alkaline conditions for concrete and soil-improvement industries.

The advantages of microbial induced carbonate mineralization for soil-stabilization and building-material industries are under extensive investigation. The pH is one of the influential parameters on the desired calcium carbonate mineralization due to the resulting textures of this mineral. Moreover, the decrease in microbial growth under the extreme alkaline environment compatible with the sustainability of concrete has been the bottleneck for an effective application of Microbial Induced Carbonate Precipitation (MICP) in the concrete industry. Microbial consortia have shown more robustness in their resistance to environmental fluctuations than pure cultures. In addition, microorganisms obtained from alkaline environments could facilitate their adaptation to extreme alkalinity. The aim of this study was to obtain urease producing bacteria (UPB) able to maintain a high MICP performance under extremely alkaline conditions compatible with concrete by adapting native microorganisms obtained from extreme environments. The growth performance, urease activity, strength of the generated biocement, and CaCO3 mineralogy were compared with the best-performer urease-producing bacteria (UPB), S. pasteurii DSMZ 33. The native bacteria presented a similar performance in growth and urease activity than S. pasteurii under extreme alkaline conditions (pH 12.5). However, the generated biocement of native Sporosarcina sp. achieved 461 % more unconfined compressive strength (UCS) and 120 % more CaCO3 content than the biocement generated by S. pasteurii DSMZ 33. The careful adaptation process performed in this study for native UPB and S. pasteurii DSMZ 33 is an interesting approach with promising and projectable results for future engineering and biotechnological applications. These results have important implications for the design of engineering solutions involving MICP. STATEMENT OF SIGNIFICANCE: A consolidated and strong biocement was generated by a native species obtained from extreme ecosystems in an effort of bioprospecting to enhance the performance of biotechnological solutions for geotechnical applications in the concrete and soil-improvement industries. Biocement generated by the native species was stronger than the generated by one of the best-described biocementation performers. This native species was able to actively growing and do perform microbial-induced-carbonate-mineralization under extreme alkalinity conditions after a careful laboratory adaptation process. The native species presented unique and differentiating traits that gave it a better adaptability and biocementation performance. The same occurs with a priceless microbial diversity inhabiting little explored and unprotected extreme ecosystems. Extreme environments house a fascinating biodiversity with potential value for ecosystem services.

Validation of Genetic Markers Associated to Oxygen Availability in Low-Grade Copper Bioleaching Systems: An Industrial Application.

Forced aeration is one of the major energy consumption factors of the bioleaching process of run-of-mine ore. The effect of aeration in the microbial community has scarcely been studied at industrial level. Leptospirillum ferriphilum is one of the most representative species of the Fe3+ producing population in this kind of systems. We analyzed the effect of oxygen availability on L. ferriphilum by growth activity and transcriptional dynamics of its two terminal oxidases (cbb3 and bd complexes) under different experimental test: culture reactor, bioleaching column, and industrial heap tests. Relatively low O2 availability triggered important changes in the microbial community composition, cell growth, microbial activity and cydAB genes transcription in all cases of study. We assessed the potential role of the terminal oxidases on the adaptation to variable aeration conditions in different lifestyles of L. ferriphilum and identified transcriptional markers associated to oxygen metabolism in an industrial system. An interesting hypothesis about the possible role of the cbb3 complex in the response to oxidative stress as well as their role as a high oxygen-affinity oxidase in L. ferriphilum is proposed and discussed. This study successfully proves the function of the cydAB genes as valid genetic markers for low-grade copper industrial bioleaching systems.

A new DNA-based model for finite field arithmetic.

A Galois field G F ( p n ) with p ≥ 2 a prime number and n ≥ 1 is a mathematical structure widely used in Cryptography and Error Correcting Codes Theory. In this paper, we propose a novel DNA-based model for arithmetic over G F ( p n ) . Our model has three main advantages over other previously described models. First, it has a flexible implementation in the laboratory that allows the realization arithmetic calculations in parallel for p ≥ 2 , while the tile assembly and the sticker models are limited to p = 2 . Second, the proposed model is less prone to error, because it is grounded on conventional Polymerase Chain Reaction (PCR) amplification and gel electrophoresis techniques. Hence, the problems associated to models such as tile-assembly and stickers, that arise when using more complex molecular techniques, such as hybridization and denaturation, are avoided. Third, it is simple to implement and requires 50 ng/µL per DNA double fragment used to develop the calculations, since the only feature of interest is the size of the DNA double strand fragments. The efficiency of our model has execution times of order O ( 1 ) and O ( n ) , for the addition and multiplication over G F ( p n ) , respectively. Furthermore, this paper provides one of the few experimental evidences of arithmetic calculations for molecular computing and validates the technical applicability of the proposed model to perform arithmetic operations over G F ( p n ) .

Validación del índice de fertilidad en endometriosis (EFI) para predecir el embarazo en una población infértil / Validation of fertility index in endometriosis (EFI) to predict pregnancy in an infertile population

Objetivo: Determinar si el Índice de Fertilidad en Endometriosis (EFI) es útil para estimar el pronóstico reproductivo en pacientes infértiles con diagnóstico y tratamiento quirúrgico de endometriosis. Diseño: Estudio de cohorte retrospectivo. Material y Métodos: Se analizaron las historias clínicas de 65 pacientes que consultaron por infertilidad entre Abril de 2011 a Septiembre de 2014 a las cuales se les realizó una videolaparoscopía diagnóstica con los mismos operadores quirúrgicos y con hallazgo de endometriosis. Se excluyeron del análisis todas aquellas pacientes que presentaban factor masculino severo, factor uterino y que realizaron tratamientos de alta complejidad. Los datos de los factores quirúrgicos para la categorización de las pacientes según el EFI fueron obtenidos a través de la visualización de videos de las laparoscopías y los factores históricos se recolectaron de las historias clínicas. Se evaluó la tasa de embarazo luego de 18 meses de seguimiento. Se subdividieron a las pacientes según los valores de EFI obtenidos en 3 grupos y se compararon las tasas acumulativas de embarazos entre dichos grupos. Resultados: La edad promedio de las pacientes fue de 33,5 años (SD=2,7). El tiempo de infertilidad promedio fue de 2,8 años (SD=1,5), y el tipo de infertilidad primaria representó el 80 % de las pacientes. La tasa total de embarazo fue de 47,7%, y según la clasificación del EFI fue 12,5% en el grupo 1, 35,7% en el grupo 2 y 69% en el grupo 3, presentando una tendencia lineal estadísticamente significativa (p=0,002). Conclusiones: Se observó que la probabilidad de embarazo espontáneo o con tratamiento de baja complejidad dentro de los 12 meses posteriores a la laparoscopía fue aumentando significativamente a medida que aumentaba la categorización del EFI. Esto nos permite considerar al EFI como una herramienta útil para estimar el pronóstico reproductivo de las pacientes infértiles con diagnóstico de endometriosis.

Objective: To determine if Endometriosis Fertility Index (EFI) is useful to estimate the reproductive outcome in infertile patients with diagnosis and surgical treatment of endometriosis. Design: Retrospective cohort study. Material and Methods: The medical records of 65 patients who consulted for infertility from April 2011 to September 2014 which underwent a diagnostic videolaparoscopy with the same surgical operators and findings of endometriosis were analyzed. All those patients with severe male factor, cervical factor and those who underwent high complexity treatments were excluded from the analysis. Data from surgical factors for categorization of patients according to EFI was obtained from videos of laparoscopy, and historical factors were collected from medical records. The pregnancy rate was evaluated after an 18- month follow-up. Patients were divided according to EFI values obtained in 3 groups, and cumulative pregnancy rates among these groups were compared. Results: The mean age of the patients was 33.5 years (SD = 2.7). The mean infertility time was 2.8 years (SD = 1.5), and primary infertility accounted for 80% of the patients. The total pregnancy rate was 47.7%, being 12.5% in group 1, 35.7% in group 2 and 69% in group 3. Conclusions: It was observed that in patients with higher EFI category, the probability of spontaneous pregnancy or low complexity treatment was increased within 12 months after laparoscopy, in a statistically significant way (p = 0.002). This allows us to validate the EFI as a useful tool to estimate the reproductive prognosis of infertile patients diagnosed with endometriosis.

Eficácia do selamento provisório de três materiais restauradores ante a solução de nitrato de prata a 50% / Effectiveness of the temporary sealing of three restorative materials to the 50% silver nitrate solution

Introdução e objetivo: Este estudo teve como objetivo analisar três materiais restauradores provisórios, Coltosol®, Cavit® W e Cimpat®,quanto a sua capacidade seladora ante a solução de nitrato de prata a50%. Material e métodos: Trinta corpos de prova foram divididos em três grupos experimentais. Cada amostra continha em seu interior um disco de antibiograma com 5 mm de diâmetro e 1 mm de espessura e,sobre o disco, 4 mm de um dos três materiais. Depois de realizadas as restaurações, os corpos de prova foram colocados em recipientes contendo nitrato de prata a 50%, sendo levados à estufa a 37ºC, na qual permaneceram por 7 dias. Decorrido esse período e comos materiais restauradores removidos, foram feitas as análises dos discos de antibiograma, que receberam escores conforme a infiltração.Resultados e conclusão: Os dados obtidos foram submetidos ao teste estatístico não-paramétrico Kruskal-Wallis, ao nível de significância de 5%. Os resultados mostraram que o Cimpat® apresentou os maioresníveis de infiltração. Já o Coltosol® e o Cavit® W não diferiram entre si (p > 0.05).

Introduction and objective: The aim of this study was to analyze three temporary restorative materials Coltosol®, Cavit® W and Cimpat®, concerning their sealing capacity to the 50% silver nitrate solution.Material and methods: 30 samples were divided into 3 experimental groups. Inside of each sample there was an antibiogram disc with 5mm of diameter and 1 mm of depth, with 4 mm of one of the three sealing materials on it. After restorations, the samples were insertedinto recipients with 50% silver nitrate solution, which were taken to theheater at 37ºC, where they remained for 7 days. Following this period and with the restorative materials removed, antibiogram disc analyses were performed, which were scored according to the leakage. Results and conclusion: Data obtained were submitted to the Kruskal-Wallisnon-parametric statistical test, with a level of significance of 5%. Results showed that Cimpat® had the greatest levels of leakage, while Coltosol® and Cavit® W did not differ between themselves (p > 0.05).