Long lasting effects of O3 and SO2 on the sensitivity to drought in two clones of Picea abies (L.) Karst.

Rooted cuttings of two Picea abies-clones were fumigated with O₃ and O₃+ SO₂ for 12 weeks. After 4 and 16 months without fumigation the treated plants showed much higher sensitivity to drought (expressed in osmotic potential of cell sap and in percent survival) than the untreated ones.     

Seasonal variations in location and population structure of endophytes in buds of Scots pine

We studied the location and distribution of a bacterial isolate, a Mycobacterium sp., in buds of Scots pine (Pinus sylvestris L.). Using a probe specific for the 16S rRNA of the Mycobacterium sp., the bacterium was found by in situ hybridization in the meristematic tissues of 40% of all bud samples examined. Because we had previously found other bacterial and fungal endophytes in the meristematic tissues of Scots pine buds, we studied their occurrence in buds during shoot development and dormancy. Using probes targeted to the 16S or 18S rRNA of the endophytes Mycobacterium sp., Methylobacterium spp., Pseudomonas spp. and Rhodotorula minuta, endophytes were found in association with growing tissues, with Methylobacterium spp. being the dominant species. Endophytes were detected in abundance before elongation or differentiation of a bud, but once a tissue was fully developed, endophytes were no longer detected. Metabolic activity of the endophytes was suppressed at the onset of, and during, dormancy of Scots pine, but recovered before the following growing season.     

Effects of Biochar and Marble mud on Mine Waste Properties to Reclaim Tailing Ponds

The effects of biochar addition in improving soil physical properties are not clearly understood in mining tailings. The objective of this study was to determine the effects of three different types of biochars, in addition to marble mud (MM) and their mixtures, on the structural stability and water retention of mine wastes in Cartagena, Spain. Biochars were produced at 500 °C from pig manure (PM), cotton (Gossipium hirsutum L.) crop residues (CR) and municipal solid waste (MSW). Biochars were added to the mine waste (MW) along with MM and a control (no amendments added). These mixtures were incubated in cores for 90 days (25 °C). PM and CR mixed with MM decreased soil bulk density (from 0 · 98 g cm^‐3 to 0 · 89 and 0 · 84 g cm⁻³, respectively). Amendments had no significant effect on total porosity whereas they increased gas diffusion by 100%, except for MSW. MM improved the plant available water from 0 · 59 to 2 · 56 cm as its combination with biochars, extremely relevant in water scarce climates. The micropores were likely replaced by mesopores when application of PM, CR, MM, and biochars + MM and they improved water retention. Total carbon (TC) and total nitrogen (TN) increased by using biochars and MM and no significant effects were assessed on aggregates. In general, MM mixed with PM and CR derived biochar improved the structural stability and exhibited a strong impact in reclaiming physical quality on mine tailings. Copyright © 2016 John Wiley & Sons, Ltd.     

Direct in vivo evidence of protective effects of grape seed procyanidin fractions and other antioxidants against ethanol-induced oxidative DNA damage in mouse brain cells

Ethanol is a principle ingredient of alcoholic beverages with potential neurotoxicity and carcinogenicity, and the ethanol-associated oxidative DNA damage in the central nervous system is well documented. The present work studied the possible protective effects of grape seed oligomer and polymer procyanidin fractions against ethanol-induced toxicity and compared these with resveratrol and other well-known antioxidants (ascorbic acid and vitamin E). By using the single cell gel electrophoresis (comet assay), a simple and sensitive technique for genotoxicity studies, the potential genotoxicity of acute and chronic ethanol administration in the different brain regions was investigated. Acute ethanol administration, at the dose of 2.5 or 5.0 g kg(-1) i.p., could induce significant DNA damage in cerebellum and hippocampus. Chronic administration of ethanol at the dose of 2.5 or 5.0 g kg-1 p.o. for 30 days could induce significant DNA damage in cerebellum, hippocampus, hypothalamus, and cortex, which could be auto-repaired at least 3 days after ethanol withdrawal. Oral administration of grape seed oligomer and polymer procyanidins and resveratrol (25, 50, and 100 mg kg(-1)) for 3 days before acute ethanol (5.0 g kg(-1), i.p.) or repeated administration of these substances together with ethanol (5.0 g kg(-1), p.o.) for 30 consecutive days could significantly inhibit DNA damage in brain cells induced by ethanol. As compared, ascorbic acid (50, 100, and 200 mg kg(-1)) and vitamin E (100, 200, and 400 mg kg(-1)) could also present protective effects on ethanol-induced DNA damage. Furthermore, the concentrations of ethanol and acetaldehyde in brain regions of the mice were detected by gas chromatography after administration of ethanol plus antioxidants. All of the results indicated that ethanol could induce region-specific oxidative DNA damage in which the cerebellum and hippocampus were more vulnerable, but intake of grape seed procyanidins or other natural antioxidants could protect the brain against ethanol-induced genotoxicity.     

Interaction of four monoterpenes contained in essential oils with model membranes: implications for their antibacterial activity

The present article reports the antimicrobial efficacy of four monoterpenes (thymol, carvacrol, p-cymene, and gamma-terpinene) against the Gram-positive bacterium Staphylococcus aureus and the Gram-negative bacterium Escherichia coli. For a better understanding of their mechanism of action, the damage caused by these four monoterpenes on biomembranes was evaluated by monitoring the release, following exposure to the compounds under study, of the water-soluble fluorescent marker carboxyfluorescein (CF) from large unilamellar vesicles (LUVs) with different lipidic composition (phosphatidylcholine, PC, phosphatidylcholine/phosphatidylserine, PC/PS, 9:1; phosphatidylcholine/stearylamine, PC/SA, 9:1). Furthermore, the interaction of these terpenes with dimyristoylphosphatidylcholine multilamellar vesicles as model membranes was monitored by means of differential scanning calorimetry (DSC) technique. Finally, the results were related also with the relative lipophilicity and water solubility of the compounds examined. We observed that thymol is considerably more toxic against S. aureus than the other three terpenes, while carvacrol and p-cymene are the most inhibitory against E. coli. Thymol and carvacrol, but not gamma-terpinene and p-cymene, caused a concentration-dependent CF leakage from all kinds of LUVs employed; in particular, thymol was more effective on PC and PC/SA LUVS than on PC/PS vesicles, while carvacrol challenge evoked a CF leakage from PC/PS LUVs similar to that induced from PC/SA LUVs, and lower than that measured with PC vesicles. Concerning DSC experiments, these four terpenes caused a decrease in Tm and (especially carvacrol and p-cymene) DeltaH values, very likely acting as substitutional impurities. Taken together, our findings lead us to speculate that the antimicrobial effect of thymol, carvacrol, p-cymene, and gamma-terpinene may result, partially at least, from a gross perturbation of the lipidic fraction of the plasmic membrane of the microorganism. In addition to being related to the physicochemical characteristics of the compounds (such as lipophilicity and water solubility), this effect seems to be dependent on the lipidic composition and net surface charge of the microbic membranes. Furthermore, the compounds might cross the cell membranes, thus penetrating into the interior of the cell and interacting with intracellular sites critical for antibacterial activity.     

A rapid multiplexed chemiluminescent immunoassay for the detection of Escherichia coli O157:H7, Yersinia enterocolitica, Salmonella typhimurium, and Listeria monocytogenes pathogen bacteria

A simple and rapid multiplexed sandwich chemiluminescent enzyme immunoassay has been developed for the simultaneous detection of Escherichia coli O157:H7, Yersinia enterocolitica, Salmonella typhimurium, and Listeria monocytogenes. To achieve the multiplexed detection of the four pathogens, a new polystyrene 96 well microtiter plate format has been designed, in which each main well contains four subwells in the bottom. The monoclonal antibodies specific for each bacteria were separately immobilized in each subwell. When the samples were added to the main wells, the bacteria able to specifically bind to the corresponding monoclonal antibody were captured in one of the four subwells. Subsequently, a mixture of peroxidase-labeled polyclonal antibodies against the four bacteria was added and the peroxidase activity of the bound polyclonal labeled antibodies in each well was measured by an enhanced luminol-based chemiluminescent cocktail using a low-light charge-coupled imaging device. The assay was simple and fast, and the limit of quantification was in the order of 104-105 CFU/mL for all bacterial species. The accuracy of the method, evaluated by comparison of the results with a conventional culturing methodology, was satisfactory, with recovery values ranging from 90 to 120%. This method can be used as a screening test to evaluate the presence of these pathogen bacteria in different foodstuffs.     

Cocoa-enriched diet enhances antioxidant enzyme activity and modulates lymphocyte composition in thymus from young rats

Cocoa is a rich source of flavonoids, mainly (-)-epicatechin, (+)-catechin, and procyanidins. This article reports the effect of continuous cocoa intake on antioxidant capacity in plasma and tissues, including lymphoid organs and liver, from young rats. Weaned Wistar rats received natural cocoa (4% or 10% food intake) for three weeks, corresponding to their infancy. Flavonoid absorption was confirmed through the quantification of epicatechin metabolites in urine. Total antioxidant capacity (TAC) and the activity of antioxidant enzymes, superoxide dismutase (SOD) and catalase, were examined. Cocoa intake enhanced TAC in all tissues especially in thymus. Moreover, thymus SOD and catalase activities were also dose-dependently increased by cocoa. It was also analyzed whether the enhanced antioxidant system in thymus could influence its cellular composition. An increase in the percentage of thymocytes in advanced development stage was found. In summary, cocoa diet enhances thymus antioxidant defenses and influences thymocyte differentiation.     

Inhibition of photophosphorylation and electron transport chain in thylakoids by lasiodiplodin, a natural product from Botryosphaeria rhodina

Four natural products were isolated from the fungus Botryosphaeria rhodina, and their effects on photosynthesis were tested. Only lasiodiplodin (1) inhibited ATP synthesis and electron flow from water to methylviologen; therefore, it acts as a Hill reaction inhibitor in freshly lysed spinach thylakoids. Photosystem I and II and partial reactions as well as ATPase were measured in the presence of 1. Three new different sites of 1 interaction and inhibition were found: one at CF1, the second in the water-splitting enzyme, and the third at the electron-transfer path between P680 and QA; these targets are different from that of the synthetic herbicides present. Electron transport chain inhibition by 1 was corroborated by fluorescence induction kinetics studies.     

Interaction of resveratrol and its trimethyl and triacetyl derivatives with biomembrane models studied by differential scanning calorimetry

The interaction of resveratrol (trans-3,5,4'-trihydroxystilbene) and two of its derivatives (3,5,4'-tri-O-methylresveratrol and 3,5,4'-tri-O-triacetylresveratrol) with biomembrane models, represented by dimyristoylphosphatidylcholine (DMPC) multilamellar vesicles (MLV), has been studied by differential scanning calorimetry (DSC). The analysis of MLV prepared in the presence of increasing molar fraction of such compounds has been carried out to reveal their maximum interaction with biomembrane models. The results from these studies have been compared with kinetic experiments results, in order to detect the entity and rate of compound absorption by the biomembrane models. The findings indicate that the compounds affected the thermotropic properties of DMPC MLV by suppressing the pretransition peak and broadening the DMPC main phase transition calorimetric peak and shifting it to lower temperatures. The order of effectiveness found was resveratrol > trimethylresveratrol > triacetylresveratrol. The kinetic experiments reveal that in an aqueous medium the absorption of resveratrol by the biomembranes models is allowed, whereas the absorption of its derivatives is hindered; in contrast when a lipophilic medium is employed, all three compounds are easily absorbed.