Variation of DNA methylation and phenotypic traits following unilateral sexual polyploidization in <Emphasis Type="Italic">Medicago</Emphasis>

Sexual hybridization is an important generator of biodiversity and a powerful breeding tool. Hybridization can also overcome ploidy barriers when it involves 2n gametes, as in the case of unilateral sexual polyploidization (USP) that has been utilized in several crops, among which alfalfa. This research was aimed at gaining insights into the effects of USP on genome methylation and on phenotypic traits in alfalfa, an important forage species. The Methylation-Sensitive Amplified Polymorphism technique was used to estimate the cytosine methylation changes occurring in a tetraploid (2n = 4x = 32) USP progeny from crosses between a diploid Medicago sativa subsp. falcata genotype that produces 2n eggs and a cultivated tetraploid Medicago sativa subsp. sativa variety. De novo methylation or demethylation in the USP progeny were observed for 13% of the detected genomic sites, indicating that methylation changes can be relevant. USP plants showed larger surface area of the leaf epidermis cells than both parents, but this did not result in larger leaf size or higher plant biomass. They displayed significant higher ovule sterility than the tetraploid parent, but normal fertility was observed in crosses with unrelated male testers. We conclude that hybridization and sexual polyploidization resulted in novel variation in terms of remodeling of the methylation landscape as well as changes in phenotypic traits in alfalfa.     

Secretion of multi-protein migratory complex induced by Toxoplasma gondii infection in macrophages involves the uPA/uPAR activation system

Toxoplasmosis is a world wide spread zoonosis caused by Toxoplasma gondii, an obligate intracellular parasite that is able to disseminate into deep tissues and cross biological barriers, reaching immunoprivileged sites such as the brain and retina. The parasite is able to infect macrophages and dendritic cells for dispersal throughout the body. However, the molecular mechanisms or outcomes of the subversion of the host cell are largely unknown. Recently our group established that metalloproteinases are involved in migration of infected macrophages. Herein, we evaluated the recruitment of host invasive machinery components in T. gondii infected murine macrophages. We showed by immunoprecipitation assays that MMP-9, CD44 TIMP-1 and uPAR were secreted as a multi-protein complex by infected macrophages. Zymographic analysis revealed that MMP-9 was present in its pro- and active form. Moreover, inhibition of uPA/uPAR pathway by PAI-1 decreased secretion of MMP-9 active forms, as well those associated to uPAR and TIMP-1, but not to CD44. Data presented here suggest that MMP-9 is secreted as a multiprotein complex by T. gondii infected macrophages, similar to that observed in metastatic cells. We further speculate that uPA/uPAR system is involved in the expression/secretion of complexes containing active MMP-9 forms.     

Determination of organic and inorganic carbon, δ13C,and nitrogen in soils containing carbonates after acid fumigation with HCl

In carbonate‐containing soils a reliable determination of organic C requires a method that effectively separates organic and inorganic C without altering the organic matter. This study was conducted to determine whether HCl vapor completely removes carbonates even in dolomite‐rich soils and to what extent a widely used acid‐fumigation method has to be modified for humus‐rich soils. Furthermore, it was tested whether HCl fumigation alters organic‐C content. Since C and N parameters are often analyzed simultaneously we also tested the influence of acid‐vapor treatment on N content and on δ¹³C of soil organic matter. We applied fumigation with 37% HCl for 8 and 32 h using 9 carbonate‐containing soil samples. Inorganic C ranged from 7 to 124 and organic C from 9 to 267 g kg^–1. The maximum contents of dolomite and calcite were 940 and 640 g kg^–1, respectively. A time of 8 h was enough to completely remove all carbonates. Neither the content nor the δ¹³C of organic C were significantly affected by fumigation. In contrast, N contents were altered by acid treatment. Based on these results and on our experience in analyzing more than 1000 soil samples, a recommended procedure for acid fumigation of carbonate‐containing soils with a wide range of organic‐ and inorganic‐C contents was derived. Samples pretreated in this way can be analyzed reliably for their organic‐C content and δ¹³C. Furthermore, N and inorganic‐C contents can be determined with a quality sufficient for many purposes.     

Computed tomography-guided fine-needle aspiration and tissue-core biopsy of bone lesions in small animals

In humans, free-hand computed tomography (CT)-guided biopsy is an accurate method to obtain a tissue sample. There are only a few reports of this technique in veterinary medicine. In the present study, 21 dogs and two cats underwent a free-hand CT-guided tissue-core biopsy (17 animals) or fine-needle aspiration (six animals) of a bone lesion. Two out of 17 tissue-core samples were also cultured. All 17 tissue-core biopsy samples were diagnostic (accuracy of 100%). Five out of six aspirates were diagnostic (accuracy of 83.3%). The overall accuracy was 95.7%. In one aspirate, cytologic quality was insufficient containing only blood. No major complications were encountered. Fourteen neoplastic, two infectious and six benign lesions were diagnosed. CT examination after intravenous contrast medium added useful information to avoid large vessels and to biopsy-viable tissue. Free-hand CT-guided tissue-core biopsy and aspiration appears to be a safe and very accurate procedure for use in the diagnosis of bone-associated diseases in small animals.     

Spin crossover in ferropericlase at high pressure: a seismologically transparent transition?

Seismic discontinuities in Earth typically arise from structural, chemical, or temperature variations with increasing depth. The pressure-induced iron spin state transition in the lower mantle may influence seismic wave velocities by changing the elasticity of iron-bearing minerals, but no seismological evidence of an anomaly exists. Inelastic x-ray scattering measurements on (Mg(0.83)Fe(0.17))O-ferropericlase at pressures across the spin transition show effects limited to the only shear moduli of the elastic tensor. This explains the absence of deviation in the aggregate seismic velocities and, thus, the lack of a one-dimensional seismic signature of the spin crossover. The spin state transition does, however, influence shear anisotropy of ferropericlase and should contribute to the seismic shear wave anisotropy of the lower mantle.     

Sequestering ability of butylated hydroxytoluene,propyl gallate,resveratrol, and vitamins C and E against ABTS,DPPH, and hydroxyl free radicals in chemical and biological systems

The antioxidant capacity of butylated hydroxytoluene (BHT; 2,6-di-tert-butyl-p-cresol), propyl gallate (3,4,5-trihydroxybenzoic acid n-propyl ester), resveratrol (trans-3,4',5-trihydroxystilbene), and vitamins C (l-ascorbic acid) and E [(+)-alpha-tocopherol] was studied in chemical and biological systems. The chemical assays evaluated the capacity of these antioxidants to sequester 2,2'-azinobis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS.) and 1,1 diphenyl-2-picrylhydrazyl (DPPH.). A new colorimetric method to determine hydroxyl radical scavenging is also described. The biological tests use the eucaryotic cells of Saccharomyces cerevisiae treated with the antioxidants in the presence of the stressing agents apomorphine, hydrogen peroxide, and paraquat dichloride (methylviologen; 1,1'-dimethyl-4,4'-bipyridinium dichloride). The results in chemical systems showed that all of the antioxidants were able to significantly inhibit the oxidation of beta-carotene by hydroxyl free radicals. The assays in yeast showed that the antioxidant activity of the tested compounds depended on the stressing agent used and the mechanism of action of the antioxidant.     

Nitrogen and phosphorous budgets during a farming cycle of the Manila clam Ruditapes philippinarum: An in situ experiment

In the Sacca di Goro lagoon a farming cycle of the Manila clam (Ruditapes philippinarum) was simulated seeding young molluscs in an unexploited sandy spot. The experimental area (2100 m²) consisted of three sectors: a control (C), almost devoid of clams (∼ 1600 m^− 2, ∼ 30 ind m^− 2), a low (L) density area (400 m², ∼ 300 ind m^− 2) and a high (H) density zone (∼ 110 m^− 2, ∼ 800 ind m^− 2). Water chemistry, external freshwater nutrient loads, molluscs filtration rates, biomass, elemental composition and nutrient recycling were analysed.Clam filtration rates and light and dark fluxes of nutrients were measured with intact core incubations. Three replicate cores (i.d. 20 cm) were collected from C, L and H in April, one month after the seeding, June, August and October 2003. External loads were calculated multiplying dissolved and particulate nutrients concentration by freshwater flow from the main lagoon tributaries. Direct excretion, filtration activity of clams and particulate matter deposition resulted in significantly higher ammonium (NH₄⁺) and soluble reactive phosphorus (SRP) effluxes to the water column at L and H. For the entire farming cycle, particulate nitrogen (PN) uptake by clams from the water column was 1.7 (C), 9.1 (L) and 16.3 (H) mol m^− 2, whilst total dissolved nitrogen (TDN) fluxes were − 0.3 (C), 1.6 (L) and 6.9 (H) mol m^− 2. Particulate phosphorus (PP) uptake from the water column was 0.1 (C), 0.6 (L) and 1.0 (H) mol m^− 2, whilst total dissolved phosphorus (TDP) efflux was 0.2 (C), 0.5 (L) and 0.8 (H) mol m^− 2. At the end of the farming cycle, harvested N as mollusc flesh was negligible for C, 0.4 mol m^− 2 for L and 1.8 mol m^− 2 for H. Harvested P as mollusc flesh was negligible for C, 0.02 mol m^− 2 for L and 0.04 mol m^− 2 for H. Farmed areas seem to have a great potential for fast coupling between sedimentation (filter feeder mediated biodeposition) and benthic recycling. At the lagoon level, mollusc farming probably attenuates the export of particulate matter to the open sea. Our results show that a minor fraction of biodeposited N (∼ 6%) and P (∼ 3%) was exported as a commercial product at the end of the farming cycle, whilst a larger fraction was incorporated in the sediments or recycled as dissolved inorganic or organic forms.     

Anti-inflammatory, antioxidant and antifungal furanosesquiterpenoids isolated from Commiphora erythraea (Ehrenb.) Engl. resin

The topical anti-inflammatory, free radical scavenging and antifungal activities of essential oils and extracts of Commiphora erythraea (Ehrenb.) Engl. resin were investigated. The hexane extract significantly inhibited oedema when applied topically in Croton oil-induced ear oedema assay in mice. The same extract showed antioxidant activity in DPPH radical scavenging assay. A bioguided separation of the hexane extract led to the isolation of furanosesquiterpenoids 1 and 2 that showed a weak antifungal activity, while compounds 3-5 resulted to be antioxidant (EC₅₀ 4.28, 2.56 and 1.08 mg/mL, respectively) and anti-inflammatory (30, 26 and 32% oedema reduction, respectively).