Development of super nanoantimicrobials combining AgCl, tetracycline and benzalkonium chloride.

In this work, we demonstrate that a simple argentometric titration is a scalable, fast, green and robust approach for producing AgCl/antibiotic hybrid antimicrobial materials. We titrated AgNO3 into tetracycline hydrochloride (TCH) aqueous solution, thus forming AgCl/TCH in a one-step procedure. Furthermore, we investigated the one-pot synthesis of triply synergistic super-nanoantimicrobials, combining an inorganic source of Ag+ ions (AgCl), a disinfecting agent (benzyl-dimethyl-hexadecyl-ammonium chloride, BAC) and a molecular antibiotic (tetracycline hydrochloride, TCH). Conventional antimicrobial tests, industrial biofilm detection protocols, and in situ IR-ATR microbial biofilm monitoring, have been adapted to understand the performance of the synthesized super-nanoantimicrobial. The resulting hybrid AgCl/BAC/TCH nanoantimicrobials are found to be synergistically active in eradicating Salmonella enterica and Lentilactobacillus parabuchneri bacteria and biofilms. This study paves the way for the development of a new class of super-efficient nanoantimicrobials that combine relatively low amounts of multiple active species into a single (nano)formulation, thus preventing the development of antimicrobial resistance towards a single active principle.

Using staining as reference for spectral imaging: Its application for the development of an analytical method to predict the presence of bacterial biofilms.

At present, although spectral imaging is known to have a great potential to provide a massive amount of valuable information, the lack of reference methods remains as one of the bottlenecks to access the full capacity of this technique. This work aims to present a staining-based reference method with digital image treatment for spectral imaging, in order to propose a fast, efficient, contactless and non-invasive analytical method to predict the presence of biofilms. Spectral images of Pseudomonasaeruginosa biofilms formed on high density polyethylene coupons were acquired in visible and near infrared (vis-NIR) range between 400 and 1000 nm. Crystal violet staining served as a biofilm indicator, allowing the bacterial cells and the extracellular matrix to be marked on the coupon. Treated digital images of the stained biofilms were used as a reference. The size and pixels of the hyperspectral and digital images were scaled and matched to each other. Intensity color thresholds were used to differentiate the pixels associate to areas containing biofilms from those ones placed in biofilm-free areas. The model facultative Gram-negative bacterium, P. aeruginosa, which can form highly irregularly shaped and heterogeneous biofilm structures, was used to enhance the strength of the method, due to its inherent difficulties. The results showed that the areas with high and low intensities were modeled with good performance, but the moderate intensity areas (with potentially weak or nascent biofilms) were quite challenging. Image processing and artificial neural networks (ANN) methods were performed to overcome the issues resulted from biofilm heterogeneity, as well as to train the spectral data for biofilm predictions.

Biochar impacts on runoff and soil erosion by water: A systematic global scale meta-analysis.

Biochar application to soil has the potential to affect soil and vegetation properties that are key for the processes of runoff and soil erosion. However, both field and pot experiments show a vast range of effects, from strong reductions to strong increases in runoff and/or soil erosion. Therefore, this study aimed to quantify and interpret the impacts of biochar on runoff and soil erosion through the first systematic meta-analysis on this topic. The developed dataset consists of 184 pairwise observations for runoff and soil erosion from 30 independent studies but 8 of which just focused on soil erosion. Overall, biochar application to soil significantly reduced runoff by 25 % and erosion by 16 %. Mitigation of soil erosion in the tropics was approximately three times stronger (30 %) than at temperate latitudes (9 %); erosion reduction in the subtropical zone was 14 %, but not significantly different from either the tropical or temperate zones. Fewer reported field observations for runoff resulted in larger confidence intervals and only the temperate latitudes showed a significant effect (i.e. a 28 % reduction). At topsoil gravimetric biochar concentrations between 0.6 % and 2.5 %, significant reductions occurred in soil erosion, with no effect at lower and higher concentrations. Biochar experiments that included a vegetation cover reduced soil erosion more than twice as much as bare soil experiments, i.e. 27 % vs 12 %, respectively. This suggests that soil infiltration, canopy interception, and soil cohesion mechanisms may have synergistic effects. Soil amended with biochar pyrolyzed at >500 °C was associated with roughly double the erosion reduction than soil amended with biochar produced at 300-500 °C, which potentially could be related to the enhancement of hydrophobicity in the latter case. Our results demonstrate substantial potential for biochar to improve ecosystem services that are affected by increased infiltration and reduced erosion, while mechanistic understanding needs to be improved.

Development of a Portable Near-Infrared Spectroscopy Tool for Detecting Freshness of Commercial Packaged Pork.

Real-time monitoring of meat quality requires fast, accurate, low-cost, and non-destructive analytical methods that can be used throughout the entire production chain, including the packaged product. The aim of this work was to evaluate the potential of a portable near-infrared (NIR) spectroscopy tool for the on-site detection of freshness of pork loin fillets in modified atmosphere packaging (MAP) stored on display counters. Pork loin slices were sealed in MAP trays under two proportions of O2/CO2/N2: High-Ox-MAP (30/40/30) and Low-Ox-MAP (5/20/75). Changes in pH, color, thiobarbituric acid reactive substances (TBARS), Warner−Bratzler shear force (WBSF), and microbiology (total viable counts, Enteriobacteriaceae, and lactic acid bacteria) were monitored over 15 days post-mortem at 4 °C. VIS-NIR spectra were collected from pork fillets before (through the film cover) and after opening the trays (directly on the meat surface) with a portable LABSPEC 5000 NIR system in diffuse reflectance mode (350−2500 nm). Quantitative NIR models by partial least squares regression (PLSR) showed a promising prediction ability for meat color (L*, a*, C*, and h*) and microbiological variables (R2VAL > 0.72 and RPDVAL > 2). In addition, qualitative models using PLS discriminant analysis obtained good accuracy (over 90%) for classifying pork samples as fresh (acceptable for consumption) or spoiled (not acceptable) based on their microbiological counts. VIS-NIR spectroscopy allows rapid evaluation of product quality and shelf life and could be used for on-site control of pork quality.

Highly Stable Core-Shell Nanocolloids: Synergy between Nano-Silver and Natural Polymers to Prevent Biofilm Formation.

Active investment in research time in the development and study of novel unconventional antimicrobials is trending for several reasons. First, it is one of the ways which might help to fight antimicrobial resistance and bacterial contamination due to uncontrolled biofilm growth. Second, minimizing harmful environmental outcomes due to the overuse of toxic chemicals is one of the highest priorities nowadays. We propose the application of two common natural compounds, chitosan and tannic acid, for the creation of a highly crosslinked polymer blend with not only intrinsic antimicrobial properties but also reducing and stabilizing powers. Thus, the fast and green synthesis of fine spherically shaped Ag nanoparticles and further study of the composition and properties of the colloids took place. A positively charged core-shell nanocomposition, with an average size in terms of the metal core of 17 ± 4 nm, was developed. Nanoantimicrobials were characterized by several spectroscopic (UV-vis and FTIR) and microscopic (transmission and scanning electron microscopies) techniques. The use of AgNPs as a core and an organic polymer blend as a shell potentially enable a synergistic long-lasting antipathogen effect. The antibiofilm potential was studied against the food-borne pathogens Salmonella enterica and Listeria monocytogenes. The antibiofilm protocol efficiency was evaluated by performing crystal violet assay and optical density measurements, direct visualization by confocal laser scanning microscopy and morphological studies by SEM. It was found that the complex nanocomposite has the ability to prevent the growth of biofilm. Further investigation for the potential application of this stable composition in food packaging will be carried out.

Catalytic synthesis of phenols with nitrous oxide.

The development of catalytic chemical processes that enable the revalorization of nitrous oxide (N2O) is an attractive strategy to alleviate the environmental threat posed by its emissions1-6. Traditionally, N2O has been considered an inert molecule, intractable for organic chemists as an oxidant or O-atom transfer reagent, owing to the harsh conditions required for its activation (>150 °C, 50‒200 bar)7-11. Here we report an insertion of N2O into a Ni‒C bond under mild conditions (room temperature, 1.5-2 bar N2O), thus delivering valuable phenols and releasing benign N2. This fundamentally distinct organometallic C‒O bond-forming step differs from the current strategies based on reductive elimination and enables an alternative catalytic approach for the conversion of aryl halides to phenols. The process was rendered catalytic by means of a bipyridine-based ligands for the Ni centre. The method is robust, mild and highly selective, able to accommodate base-sensitive functionalities as well as permitting phenol synthesis from densely functionalized aryl halides. Although this protocol does not provide a solution to the mitigation of N2O emissions, it represents a reactivity blueprint for the mild revalorization of abundant N2O as an O source.

Gold-Catalyzed Reaction of Propargyl Esters and Alkynylsilanes: Synthesis of Vinylallene Derivatives through a Twofold 1,2-Rearrangement.

The reaction of propargyl esters with alkynylsilanes under gold catalysis provides vinylallene derivatives through consecutive [1,2]-acyloxy/[1,2]-silyl rearrangements. Good yields, full atom-economy, broad substrate scope, easy scale-up and low catalyst loadings are salient features of this novel transformation. Density Functional Theory (DFT) calculations suggest a reaction mechanism involving initial [1,2]-acyloxy rearrangement to generate a gold vinylcarbene intermediate which upon regioselective attack of the alkynylsilane affords a vinyl cation which undergoes a type II-dyotropic rearrangement involving the silyl group and the metal fragment. Preliminary results on the enantioselective version of this transformation are also disclosed.

Physicochemical, sensory and microbiological characterization of Asturian Chorizo, a traditional fermented sausage manufactured in Northern Spain.

Industrial standardization of fermented meat products requires starter culturesto avoid random variations in the initial microbiota of food matrix. This allows to homogenize production batches regarding sensory and physicochemical characteristics. Also, starters contribute to assure safety, as they compete with pathogens or spoilage species, facilitating pH reduction and secretion of inhibitors. Asturian Chorizo is a traditional fermented sausage from Northern Spain, still produced in a traditional way, without starters. This work describes its characterization at the sensory, physicochemical and microbiological levels. In contrast to other fermented sausages, Asturian Chorizo microbiota is represented mainly by Lactobacillus plantarum, with secondary contributions from Lb. sakei and Lb. futsai. This results may path the way towards development of specific starter cultures for this product, avoiding the loss of the original characteristics of the fermented product in a traditional way, as it would happen in the case of using industrial processes with conventional starters.

Trapping para-Quinone Methide Intermediates with Ferrocene: Synthesis and Preliminary Biological Evaluation of New Phenol-Ferrocene Conjugates.

The reaction of para-hydroxybenzyl alcohols with ferrocene in the presence of a catalytic amount of InCl3 provided ferrocenyl phenol derivatives, an interesting class of organometallic compounds with potential applications in medicinal chemistry. This transformation exhibited a reasonable substrate scope delivering the desired products in synthetically useful yields. Evidence of involvement of a para-quinone methide intermediate in this coupling process was also provided. Preliminary biological evaluation demonstrated that some of the ferrocene derivatives available by this methodology exhibit significant cytotoxicity against several cancer cell lines with IC50 values within the range of 1.07⁻4.89 µM.

Synthesis of Functionalized Ferrocene Derivatives from Donor-Acceptor Cyclopropanes.

The TfOH-catalyzed reaction of aryl-substituted donor-acceptor cyclopropane derivatives with ferrocene provided new functionalized ferrocene derivatives. This process exhibited a reasonable scope with respect to the cyclopropane component affording the corresponding alkylated ferrocene derivatives in useful yields. An alternative approach to these functionalized ferrocene derivatives involving arylation of a ferrocene-decorated donor-acceptor cyclopropane derivative is also disclosed. Both processes rely on a facile regioselective ring-opening of the donor-acceptor cyclopropane.

Recent Developments in Coinage Metal Catalyzed Transformations of Stabilized Vinyldiazo Compounds: Beyond Carbenic Pathways.

Transition metal-catalyzed transformations of vinyldiazo compounds have become a versatile tool in organic synthesis. Although several transition metals have been investigated for this purpose, this field has been mainly dominated by dirhodium catalysts. Remarkable levels of chemo-, regio-, diastereo- and enantioselectivity have been reached in some of these rhodium-catalyzed transformations. In the last few years coinage metals have also emerged as useful catalysts in transformations involving vinyldiazo compounds. In some cases, highly efficient catalyst-dependent protocols arising from divergent mechanistic pathways have been reported. In this Personal Account, we aim to showcase recent advances in metal coinage catalyzed transformations of vinyldiazoacetates, an exciting field of research to which our group has actively contributed in the last few years.

Fire-induced pine woodland to shrubland transitions in Southern Europe may promote shifts in soil fertility.

Since the mid of the last century, fire recurrence has increased in the Iberian Peninsula and in the overall Mediterranean basin due to changes in land use and climate. The warmer and drier climate projected for this region will further increase the risk of wildfire occurrence and recurrence. Although the impact of wildfires on soil nutrient content in this region has been extensively studied, still few works have assessed this impact on the basis of fire recurrence. This study assesses the changes in soil organic C and nutrient status of mineral soils in two Southern European areas, Várzea (Northern Portugal) and Valencia (Eastern Spain), affected by different levels of fire recurrence and where short fire intervals have promoted a transition from pine woodlands to shrublands. At the short-term (<1year), the amount of soil organic matter was higher in burned than in unburned soils while its quality (represented as labile to total organic matter) was actually lower. In any case, total and labile soil organic matter showed decreasing trends with increasing fire recurrence (one to four fires). At the long-term (>5years), a decline in overall soil fertility with fire recurrence was also observed, with a drop between pine woodlands (one fire) and shrublands (two and three fires), particularly in the soil microsites between shrubs. Our results suggest that the current trend of increasing fire recurrence in Southern Europe may result in losses or alterations of soil organic matter, particularly when fire promotes a transition from pine woodland to shrubland. The results also point to labile organic matter fractions in the intershrub spaces as potential early warning indicators for shifts in soil fertility in response to fire recurrence.

The Sp1/Egr1-tandem repeat polymorphism in the 5-lipoxygenase gene promoter is not associated with late onset Alzheimer disease.

Arachidonate 5-lipoxygenase plays an important role in the synthesis of leukotrienes. Leukotrienes are inflammatory mediators, and inflammation has been implicated in the pathogenesis of Alzheimer disease. A polymorphism in the ALOX5 promoter consisting on 3 to 6 tandem-repeats of a Sp1/Egr1 binding motif (GGGCGG)n, has been related with the amount of gene expression. To verify the association between this polymorphism and the risk for late-onset Alzheimer disease we genotyped a total of 291 patients (mean age 74+/-7 y) and 300 controls (mean age 73+/-8 y). We found alleles of 3 to 6 repeats, and allele and genotype frequencies did not differ between patients and controls. These frequencies did not differ between patients according to the APOE genotype (epsilon 34 + epsilon 44 vs. epsilon 23 + epsilon 33). Together, our results indicate that the Sp1/Egr1-repeat polymorphism in the ALOX5 promoter is not a genetic marker for the risk of developing late-onset Alzheimer disease.

Role of the CDKN1A/p21, CDKN1C/p57, and CDKN2A/p16 genes in the risk of atherosclerosis and myocardial infarction.

Atherosclerosis is characterized by excessive proliferation of neointimal leukocytes and vascular smooth muscle cells (VSMCs). In mice, the manipulation of cell cycle inhibitors such as CDKN1B (p27) and CDKN1A (p21) modifies the risk of developing atherosclerosis. In humans, CDKN1A, CDKN1B and CDKN1C (p57) are differentially expressed in normal versus atherosclerotic vessels. A DNA-polymorphism within the CDKN1B promoter has been associated with myocardial infarction (MI). In the present study, we analyzed the effect of CDKN1A, CDKN1C and CDKN2A (p16) polymorphisms on MI-risk. A total of 316 patients (all male, < 55 years) and 434 controls were genotyped, and the allele and genotype frequencies were compared between the two groups. Two CDKN1C polymorphisms, a promoter GT-repeat and a variable number of repeats of the amino acid PAPA-motif, were associated with MI. The presence of two alleles < or = 11-repeats (9/11, 10/11 and 11/11 genotypes) was significantly less frequent among patients (p < 0.001). This difference was also significant when analyzing the subpopulation of smokers (p = 0.004), suggesting a protective role for these low-repeat genotypes (OR = 0.49, 95%CI = 0.32-0.73). The PAPA-BB homozygotes were significantly less frequent in patients, but this could be attributed to a linkage disequilibrium between the 11-repeats and B alleles. No significantly different frequencies between patients and controls for the four CDKN1A (-1026A/G, -754G/C, -369G/C and Ser31Arg) and the three CDKN2A (-523 G/A, +22 G/A and Ala148Thr) polymorphisms was found. In conclusion, we provide here genetic evidence for the association between DNA-variants in the CDKN1C/p57 gene and the risk of atherosclerosis and MI.

The NOS3 (27-bp repeat, intron 4) polymorphism is associated with susceptibility to osteomyelitis.

Cytokines generate nitric oxide (NO) in osteoblasts and neutrophils through the induction of NO synthase isoforms, endothelial (NOS3) and inducible (NOS2), thereby producing bone loss. In osteomyelitis (OM), a chronic infection of the bone, homozygosity for the NOS3 (27-bp repeat, intron 4 polymorphism) 4 allele was significantly more frequent among the 80 patients than in 300 healthy controls (p=0.044). No significant differences were found for other polymorphisms of the NOS genes such as NOS3, the promoter (-786T/C), and the missense change (E298D) in exon 7, and for NOS2, the G/A substitution at position 37498 in exon 22, the (CCTTT)(n), and (TAAA)(n) micro-satellites and the -954G/C in the promoter. Serum NO levels were significantly higher only in the OM patients homozygous for the NOS3 (27-bp repeat, intron 4 polymorphism) 4 allele, compared to controls. In the presence of bacteria or bacterial products, the neutrophils of these patients produced more NO. However, immunolabelling of osteoblasts for NOS3 in biopsy tissues did not correlate with the carriage of a determined NOS polymorphism but with the presence of bone inflammation. This is the first report of an association between a NOS3 polymorphism and the risk of developing OM.

Myocyte enhancing factor-2A in Alzheimer's disease: genetic analysis and association with MEF2A-polymorphisms.

Polymorphisms at different genes have been proposed as determinants of the risk for developing late-onset Alzheimer's disease (LOAD). Among the several candidate genes are those that encode proteins involved in neuronal degeneration/survival. Studies of primary neuronal cultures supported that members of the myocyte enhancing factor-2 (MEF2) family of transcription factors have an anti-apoptotic effect, regulating the expression of proteins involved in neuronal survival and differentiation. We analysed the MEF2A gene in a total of 357 patients (mean age 72 years, range 60-97 years). Among others, a Pro279Leu in exon 8 and a polyglutamine (CAG) repeat polymorphisms in exon 12 were found. These variants were also genotyped in 495 healthy controls (>50 years old), and the frequencies were statistically compared. Eight patients were 279L (six P/L and two L/L), compared to only one control (2% vs. 0.2%; p=0.004, OR=11.32). There was a significantly higher frequency of 279L-carriers among APOE epsilon4+ (7/154=4.5%), compared to epsilon4- (1/203) (p=0.02). In conclusion, our work suggests that the variation at the MEF2A gene could be involved in the risk of developing LOAD. Because MEF2 has been related with neuronal survival, and the 279L allele has been related with a reduction in the transcriptional activation activity of MEF2A, the effect of this allele could be mediated through a down-regulation of antiapoptotic genes.

Mitochondrial DNA haplogroups in Spanish patients with hypertrophic cardiomyopathy.

Mutations in mtDNA have been implicated in the development of hypertrophic cardiomyopathy (HCM), including cases from families with a maternal transmission. Alleles at several polymorphic sites in mtDNA define different haplogroups and some of these haplogroups have been involved in the risk of developing several diseases in which mitochondria should be involved. We analysed the association between the nine common European haplogroups and HCM. A total of 130 Spanish patients and 300 healthy controls were genotyped for eight mitochondrial single nucleotide polymorphisms (SNPs) through polymerase chain reaction followed by digestion with a restriction enzyme (PCR-RFLP). We compared the frequencies of these polymorphisms and mitochondrial haplogroups between patients and controls. Haplogroup T, specifically defined by 13368A, was significantly involved in the risk of developing HCM in our population (p=0.007; OR=2.42; 95% CI=1.25-4.67). Our data suggest that the genetic variation at the mitochondrial genome could significantly contribute to the risk for HCM.

ABCA1 polymorphisms and prognosis after myocardial infarction in young patients.

High-density cholesterol (HDL) levels are affected by genetic influences and certain behaviors. Low levels of HDL-C are considered as an independent risk factor for premature coronary heart disease. In patients with Tangier disease, characterised by low HDL levels, mutations in the ATP binding cassette transporter have been described. We have analysed three polymorphisms of the ABCA1 gene (-477C/T, R219 K, and I883M) in a cohort of young male survivors of myocardial infarction in order to know their influence in long-term prognosis. In premature heart disease, knowing prognosis factors is specially relevant.

A single-nucleotide polymorphism in the human p27kip1 gene (-838C>A) affects basal promoter activity and the risk of myocardial infarction.

BACKGROUND: Excessive proliferation of vascular smooth muscle cells and leukocytes within the artery wall is a major event in the development of atherosclerosis. The growth suppressor p27kip1 associates with several cyclin-dependent kinase/cyclin complexes, thereby abrogating their capacity to induce progression through the cell cycle. Recent studies have implicated p27kip1 in the control of neointimal hyperplasia. For instance, p27kip1 ablation in apolipoprotein-E-null mice enhanced arterial cell proliferation and accelerated atherogenesis induced by dietary cholesterol. Therefore, p27kip1 is a candidate gene to modify the risk of developing atherosclerosis and associated ischaemic events (i.e., myocardial infarction and stroke). RESULTS: In this study we found three common single-nucleotide polymorphisms in the human p27kip1 gene (+326T>G [V109G], -79C>T, and -838C>A). The frequency of -838A carriers was significantly increased in myocardial infarction patients compared to healthy controls (odds ratio [OR] = 1.73, 95% confidence interval [95%CI] = 1.12-2.70). In addition, luciferase reporter constructs driven by the human p27kip1 gene promoter containing A at position -838 had decreased basal transcriptional activity when transiently transfected in Jurkat cells, compared with constructs bearing C in -838 (P = 0.04). CONCLUSIONS: These data suggest that -838A is associated with reduced p27kip1 promoter activity and increased risk of myocardial infarction.