Functionalization and Characterization of Magnetic Nanoparticles for the Detection of Ferritin Accumulation in Alzheimer's Disease

Early diagnosis in Alzheimer's disease (AD), prior to the appearance of marked clinical symptoms, is critical to prevent irreversible neuronal damage and neural malfunction that lead to dementia and death. Therefore, there is an urgent need to generate new contrast agents which reveal by a noninvasive method the presence of some of the pathological signs of AD. In the present study, we demonstrate for the first time a new nanoconjugate composed of magnetic nanoparticles bound to an antiferritin antibody, which has been developed based on the existence of iron deposits and high levels of the ferritin protein present in areas with a high accumulation of amyloid plaques (particularly the subiculum in the hippocampal area) in the brain of a transgenic mouse model with five familial AD mutations. Both in vitro and after intravenous injection, functionalized magnetic nanoparticles were able to recognize and bind specifically to the ferritin protein accumulated in the subiculum area of the AD transgenic mice.Fil: Fernández Cabada, Tamara. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Politécnica de Madrid; EspañaFil: Martínez Serrano, Alberto. Consejo Superior de Investigaciones Científicas; España. Universidad Autónoma de Madrid; EspañaFil: Cussó, Lorena. Instituto de Investigacion Sanitaria Gregorio Marañón; España. Universidad Carlos III de Madrid; España. Centro de Investigación Biomédica en Red de Salud Mental; EspañaFil: Desco, Manuel. Instituto de Investigacion Sanitaria Gregorio Marañón; España. Centro de Investigación Biomédica en Red de Salud Mental; España. Universidad Carlos III de Madrid; EspañaFil: Ramos Gómez, Milagros. Universidad Politécnica de Madrid; Españ

Zeolite/iron oxide composite as sorbent for magnetic solid-phase extraction of benzene, toluene, ethylbenzene and xylenes from water samples prior to gas chromatography-mass spectrometry

This study reports a new composite based on ZSM-5 zeolite decorated with iron oxide magnetic nanoparticles as a valuable sorbent for magnetic solid-phase extraction (MSPE). A proposal is made to determine benzene, toluene, ethylbenzene and xylenes (BTEX) as model analytes in water samples using gas chromatography-mass spectrometry. A two-step multivariate optimization strategy, using Plackett⬜Burman and circumscribed central composite designs, was employed to optimize experimental parameters affecting MSPE. The method was evaluated under optimized extraction conditions (i.e., amount of sorbent, 138 mg; extraction time, 11 min; sample pH, pH of water (i.e., 5.5⬜6.5); eluent solvent volume, 0.5 mL; and elution time, 5 min), obtaining a linear response from 1 to 100 μg L↙1 for benzene; from 10 to 100 μg L↙1 for toluene, ethylbenzene and o-xylene; and from 10 to 75 μg L↙1 for m,p-xylene. The repeatability of the proposed method was evaluated at a 40 μg L↙1 spiking level and coefficients of variation ranged between 8 and 11% (n = 5). Limits of detection were found to be 0.3 μg L↙1 for benzene and 3 μg L↙1 for the other analytes. These values satisfy the current normative of the Environmental Protection Agency and European Union for BTEX content in waters for human consumption. Finally, drinking water, wastewater and river water were selected as real water samples to assess the applicability of the method. Relative recoveries varied between 85% and 114% showing negligible matrix effects.The authors would like to thank the Spanish Ministry of Science and Innovation (project no. CTQ2011-23968), Generalitat Valenciana (Spain) (projects nos. GVA/2014/096 and PROMETEO/2013/038) for the financial support. E. Fernández also thanks Ministry of Education for her FPU grant (FPU13/03125)

Back to soil: Awakening the production of cryptic antibiotics in Streptomyces

Most of the antibiotic biosynthetic gene clusters (BGCs) in Streptomyces are not expressed under laboratory conditions, however these clusters are maintained in the genomes, therefore indicating that they must play important roles in adaptation and survival within their ecological niches. Understanding the global regulation patterns that affect transcription of antibiotic BGCs in soil conditions versus laboratory conditions allows us to identify novel routes for up- or down- regulation in order to trigger expression of these BGCs and therefore production of these antibiotics under controlled laboratory and fermentation conditions. Cryptic pathways represent an untapped resource in terms of new metabolites that could be very useful in the clinic if we can awaken their expression and production. We are using RNA sequencing to analyse global transcriptional patterns of S. venezuelae and S. rimosus grown in sterile soil, non-sterile soil and standard laboratory solid medium, as well as using an unbiased metabolomics approach to identify compounds produced in soils. These results will enable the identification of global regulatory pathways that activate cryptic clusters in nature and lead to the production of these compounds in soil. This will enable us to generate genetically modified strains engineered to increase the yield of both well characterised and cryptic clusters which encode potentially novel compounds under industrial fermentation conditions. It is worth noting that the transcriptional data may also help shed light on the complex regulation of the life cycle of these Streptomyces strains in an ecologically relevant environment

Actividades antimicrobianas derivadas de fagos y su aplicación como bioconservantes alimentarios

Comunicación presentada en el XXII Congreso Nacional SEM 2009, celebrado en Almeria del 21 al 24 de septiembre de 2009.Staphylococcus aureus es un importante patógeno asociados a intoxicaciones de tipo alimentario, y a su vez, uno de los principales agentes causantes de la mastitis en el ganado vacuno.Peer reviewe

Synergy between the phage endolysin LysH5 and nisin to kill Staphylococcus aureus in pasteurized milk

Phage-encoded endolysins are recently considered as new biocontrol tools to inhibit pathogens in food. In this work, we have studied the ionic requirements for optimal lytic activity of LysH5, the endolysin encoded by the staphylococcal bacteriophage phi-SauS-IPLA88. LysH5 activity was inhibited by the presence of Mn++ and Zn++ and enhanced by Ca++, Mg++ and NaCl. When LysH5 was combined with nisin, a bacteriocin currently used as a biopreservative in food, a strong synergistic effect was observed. The Minimum Inhibitory Concentrations of nisin and LysH5 were reduced 64- and 16-fold, respectively, as determined in checkerboard microtitre tests. In addition, nisin enhanced 8-fold the lytic activity of LysH5 on cell suspensions. The synergy observed in vitro was confirmed in challenge assays in pasteurized milk contaminated with S. aureus Sa9. Clearance of the pathogen was only achieved by the combined activity of both antimicrobials. As far as we know, this is the first study that exploits the possibilities of hurdle technology combining a phage-encoded endolysin and the bacteriocin nisin for efficient S. aureus inhibition in milk.This research study was supported by grant AGL2009-13144-C02-01 from the Ministry of Science and Innovation of Spain and IB08-052 from FICYT (Regional Government of Asturias).Peer reviewe

Natural product discovery in soil actinomycetes: unlocking their potential within an ecological context

Natural products (NPs) produced by bacteria, particularly soil actinomycetes, often possess diverse bioactivities and play a crucial role in human health, agriculture, and biotechnology. Soil actinomycete genomes contain a vast number of predicted biosynthetic gene clusters (BGCs) yet to be exploited. Understanding the factors governing NP production in an ecological context and activating cryptic and silent BGCs in soil actinomycetes will provide researchers with a wealth of molecules with potential novel applications. Here, we highlight recent advances in NP discovery strategies employing ecology-inspired approaches and discuss the importance of understanding the environmental signals responsible for activation of NP production, particularly in a soil microbial community context, as well as the challenges that remain

Rapid determination of hydrophilic phenols in olive oil by vortex-assisted reversed-phase dispersive liquid-liquid microextraction and screen-printed carbon electrodes

A novel approach is presented to determine hydrophilic phenols in olive oil samples, employing vortex-assisted reversed-phase dispersive liquid-liquid microextraction (RP-DLLME) for sample preparation and screen-printed carbon electrodes for voltammetric analysis. The oxidation of oleuropein, hydroxytyrosol, caffeic acid, ferulic acid and tyrosol was investigated, being caffeic acid and tyrosol selected for quantification. A matrix-matching calibration using sunflower oil as analyte-free sample diluted with hexane was employed to compensate matrix effects. Samples were analyzed under optimized RP-DLLME conditions, i.e., extractant phase, 1 M HCl; extractant volume, 100 µL; extraction time, 2 min; centrifugation time, 10 min; centrifugation speed, 4000 rpm. The working range showed a good linearity between 0.075 and 2.5 mg L−1 (r = 0.998, N = 7) for caffeic acid, and between 0.075 and 3 mg L−1 (r = 0.999, N = 8) for tyrosol. The methodological limit of detection was empirically established at 0.022 mg L−1 for both analytes, which is significantly lower than average contents found in olive oil samples. The repeatability was evaluated at two different spiking levels (i.e., 0.5 mg L−1 and 2 mg L−1) and coefficients of variation ranged from 8% to 11% (n = 5). The applicability of the proposed method was tested in olive oil samples of different quality (i.e., refined olive oil, virgin olive oil and extra virgin olive oil). Relative recoveries varied between 83% and 108% showing negligible matrix effects. Finally, fifteen samples were analyzed by the proposed method and a high correlation with the traditional Folin-Ciocalteu spectrophotometric method was obtained. Thereafter, the concentrations of the fifteen oil samples were employed as input variables in linear discriminant analysis in order to distinguish between olive oils of different quality.The authors would like to thank Spanish Ministry of Science and Innovation (Project no. CTQ2011-23968), Spanish Ministry of Economy, Industry and Competitiveness (Project no. CTQ2016-79991-R, AEI/FEDER, UE) and Generalitat Valenciana (Spain) (Projects n. GVA/2014/096 and PROMETEO/2013/038) for the financial support. E. Fernández also thanks Spanish Ministry of Education for her FPU grant (FPU13/03125)

Hydrophilic magnetic ionic liquid for magnetic headspace single-drop microextraction of chlorobenzenes prior to thermal desorption-gas chromatography-mass spectrometry

A new, fast, easy to handle, and environmentally friendly magnetic headspace single-drop microextraction (Mag-HS-SDME) based on a magnetic ionic liquid (MIL) as an extractant solvent is presented. A small drop of the MIL 1-ethyl-3-methylimidazolium tetraisothiocyanatocobaltate(II) ([Emim]2[Co(NCS)4]) is located on one end of a small neodymium magnet to extract nine chlorobenzenes (1,2-dichlorobenzene, 1,3-dichlorobenzene, 1,4-dichlorobenzene, 1,2,3-trichlorobenzene, 1,2,4-trichlorobenzene, 1,3,5-trichlorobenzene, 1,2,3,4-tetrachlorobenzene, 1,2,4,5-tetrachlorobenzene, and pentachlorobenzene) as model analytes from water samples prior to thermal desorption-gas chromatography-mass spectrometry determination. A multivariate optimization strategy was employed to optimize experimental parameters affecting Mag-HS-SDME. The method was evaluated under optimized extraction conditions (i.e., sample volume, 20 mL; MIL volume, 1 μL; extraction time, 10 min; stirring speed, 1500 rpm; and ionic strength, 15% NaCl (w/v)), obtaining a linear response from 0.05 to 5 μg L−1 for all analytes. The repeatability of the proposed method was evaluated at 0.7 and 3 μg L−1 spiking levels and coefficients of variation ranged between 3 and 18% (n = 3). Limits of detection were in the order of nanograms per liter ranging from 4 ng L−1 for 1,4-dichlorobenzene and 1,2,3,4-tetrachlorobenzene to 8 ng L−1 for 1,2,4,5-tetrachlorobenzene. Finally, tap water, pond water, and wastewater were selected as real water samples to assess the applicability of the method. Relative recoveries varied between 82 and 114% showing negligible matrix effects.The authors would like to thank Vicerrectorado de Investigación y Transferencia del Conocimiento of the University of Alicante (UAUSTI16-04), Generalitat Valenciana (project nos. GVA/2014/096 and PROMETEO/2013/038), and Ministerio de Economía, Industria y Competitividad (project no. CTQ2016-79991-R, AEI/FEDER, UE) for the financial support. E. Fernández thanks Ministerio de Educación for her FPU grant (FPU13/03125)

Impacto del componente servicio de alimentación sobre la relación del estado de nutrición y la calidad de la dieta en escolares de Escuelas de Tiempo Completo en una zona rural del Estado de Hidalgo, México

Introduction: The objective of the study is to evaluate the impact of the food service component on the nutritional status of schoolchildren who have the Full Time Schools Program. Material and methods: Analytical, prospective cross-sectional study in 312 first and second grade primary school students, one group of students had the Food Service Component (CSA), while the other did not (NCSA). Body Mass Index (BMI), body fat percentage and waist circumference were measured. Energy and macronutrient intake was evaluated from the frequency of weekly food consumption and intake during school hours (Lunchbox inventory and weight record and measurements). Descriptive statistics and Mann-Whithney U, Chi square or Fisher's Exact Test were applied to compare groups. Results: Mean age of 7.1 ± 0.6 years, 48.4% benefited from CSA and 51.6% NCSA. The population presented 1.3% underweight, 69.6% normal and 29.2% overweight evaluated by BMI; 11.5% elevated body fat and 13% cardiometabolic risk. The NCSA group had higher percentages of excess weight, elevated body fat, and cardiovascular risk (p <0.005). By frequency of weekly consumption, men with NCSA reported higher consumption of carbohydrates (1,499.9 kcal), proteins (377.0 kcal) and lipids (1,154.6 kcal) (p <0.005); During school hours, women with NCSA showed the highest lipid intake (216.0 kcal). According to the adequacy, in the frequency of weekly consumption the groups with CSA and NCSA was above the recommendation (68.9% and 75.2% respectively) (p <0.005); In school hours the intake was also above the recommendation of 30% both in schoolchildren with CSA (70.2%) and NCSA (95.7%) (p <0.005). Conclusions: The intake of energy and macronutrients exceeds the recommended intake of all schoolchildren, men are the most affected with excess body weight, elevated body fat and cardiovascular risk. The nutritional status of the CSA beneficiary schoolchildren improves with the quality of the diet compared to NCSA schoolchildren.Introducción: El objetivo del estudio es la evaluación del impacto del componente de servicio de alimentación sobre el estado nutricional de escolares que cuentan con el Programa de Escuelas de Tiempo Completo. Material y métodos: Estudio transversal analítico, prospectivo en 312 escolares de primer y segundo grado de nivel primaria. Un grupo contó con el Componente de Servicio de Alimentación (CSA), el otro no (NCSA). Se midió el Índice de Masa Corporal (IMC), porcentaje de grasa corporal y circunferencia de cintura. Se evaluó la ingesta de energía y macronutrientes con frecuencia de consumo de alimento semanal y la ingesta en el horario escolar (Inventario de lonchera y registro de pesos y medidas). Se aplicó estadística descriptiva y U de Mann-Whithney, Chi cuadrada o Prueba Exacta de Fisher para comparar grupos. Resultados: Edad media de 7,1 ± 0,6 años, 48,4% fue beneficiado con CSA y 51,6% NCSA. La población presentó 1,3% en bajo peso, 69,6% normal y 29,2% exceso de peso evaluado por IMC; 11,5% grasa corporal evaluada por impedancia bioeléctrica y 13% riesgo cardiometabólico. El grupo NCSA tuvo mayores porcentajes de exceso de peso, grasa corporal elevada y riesgo cardiovascular (p<0,05). Por frecuencia de consumo semanal, los hombres con NCSA reportaron mayor consumo de hidratos de carbono (1.499,9 kcal), proteínas (377,0 kcal) y lípidos (1.154,6 kcal) (p<0,05); en el horario escolar las mujeres con NCSA mostraron la mayor ingesta de lípidos (216,0 kcal). De acuerdo a la adecuación en la frecuencia de consumo semanal, los grupos con CSA y NCSA fue por arriba de la recomendación (68,9% y 75,2% respectivamente) (p<0,05); en el horario escolar la ingesta también fue por arriba de la recomendación del 30% tanto en escolares con CSA (70,2%) y NCSA (95,7%) (p<0,05). Conclusiones: La ingesta de energía y macronutrimentos exceden de la ingesta recomendada de todos los escolares, los hombres son los más afectados con exceso de peso, grasa corporal elevada y riesgo cardiovascular. El estado nutricional de los escolares beneficiados CSA mejora con la calidad de la dieta en comparación con escolares NCSA

'Artilysation' of endolysin λSa2lys strongly improves its enzymatic and antibacterial activity against streptococci

Endolysins constitute a promising class of antibacterials against Gram-positive bacteria. Recently, endolysins have been engineered with selected peptides to obtain a new generation of lytic proteins, Artilysins, with specific activity against Gram-negative bacteria. Here, we demonstrate that artilysation can also be used to enhance the antibacterial activity of endolysins against Gram-positive bacteria and to reduce the dependence on external conditions. Art-240, a chimeric protein of the anti-streptococcal endolysin λSa2lys and the polycationic peptide PCNP, shows a similar species specificity as the parental endolysin, but the bactericidal activity against streptococci increases and is less affected by elevated NaCl concentrations and pH variations. Time-kill experiments and time-lapse microscopy demonstrate that the killing rate of Art-240 is approximately two-fold higher compared to wildtype endolysin λSa2lys, with a reduction in viable bacteria of 3 log units after 10 min. In addition, lower doses of Art-240 are required to achieve the same bactericidal effect.This research study was supported by grants AGL2012-40194-C02-01 (Ministry of Science and Innovation, Spain), FEDER founds and GRUPIN14-139 (Program of Science, Technology and Innovation 2013–2017, Principado de Asturias, Spain), bacteriophage network FAGOMA and research grant 1.5.171.15N of the Research Foundation – Flanders (FWO). DG was a fellow of the Ministry of Science and Innovation, Spain. LR-R was a FWO Pegasus Marie Curie Fellow. PG, BM, RL and AR are members of the FWO Vlaanderen funded “Phagebiotics” research community (WO.016.14).Peer Reviewe