Heat treatments and expansin gene expression in strawberry fruit

Heat treatments have been applied in fruit postharvest technology for insect disinfestations, decay control, ripening delay and modification of fruit responses to other stresses. Heat treatment affects several aspects of fruit ripening, such as ethylene production and cell wall degradation probably through changes in gene expression and protein synthesis. In this paper, strawberries (Fragaria × ananassa Duch., cv Camarosa) at 50–75% red stage of ripening were heat-treated at 45 °C during 3 h in an air oven and then stored at 20 °C for 0, 4, 18, 24 and 48 h. Fruit firmness was determined and the expression of a set of expansin genes (FaEXP1, FaEXP2, FaEXP4, FaEXP5 and FaEXP6) was analyzed. The firmness of treated fruit was higher than that of control fruit 24 h after treatment, though the differences disappeared after 48 h at 20 °C. The analysis by northern-blot indicated that heat treatments affected differently the expression of expansin genes. The expression of FaEXP1, FaEXP2 and FaEXP6 was lower in treated fruit during the following 24 h post-treatment. The lower expression of these expansin genes could contribute to delay softening after heat treatment.     

Characterisation of the first <Emphasis Type="Italic">Stemphylium vesicarium</Emphasis> isolates resistant to strobilurins in Italian pear orchards

Up to 2005 the sensitivity of Stemphylium vesicarium (Wallr.) Simm., the causal agent of pear brown spot, to the strobilurin fungicides kresoxim-methyl, trifloxystrobin and pyraclostrobin was still comparable with baseline values associated with good efficacy in the field. During 2006, the first resistant isolates were detected in two commercial pear orchards in the Emilia-Romagna region (Italy), one of which was affected by considerable control failure linked to strobilurin treatments as demonstrated in a field trial. In vitro sensitivity tests with 0.5 mg l⁻¹ of kresoxim-methyl, trifloxystrobin and pyraclostrobin showed that in the population collected in the orchard with control failure the conidial germination was greater than 90% compared to an untreated control both in 2006 and in 2007, i.e. 1 year after the suspension of strobilurin applications. In the other orchard, where only a few symptomatic fruits were found and the strobilurins were still in use, the conidial germination was lower, about 50% in 2006 and 25% in 2007. The molecular analysis of mitochondrial cytochrome b gene of some monospore isolates with different levels of sensitivity confirmed the presence of the mutation causing G143A substitution in all the resistant isolates. In conclusion, both in vitro tests and molecular analysis confirmed the first occurrence of Stemphylium vesicarium resistance to all strobilurin fungicides tested.     

Reversible changes in cell morphology due to cytoskeletal rearrangements measured in real-time by QCM-D

The mechanical properties and responses of cells to external stimuli (including drugs) are closely connected to important phenomena such as cell spreading, motility, activity, and potentially even differentiation. Here, reversible changes in the viscoelastic properties of surface-attached fibroblasts were induced by the cytoskeleton-perturbing agent cytochalasin D, and studied in real-time by the quartz crystal microbalance with dissipation (QCM-D) technique. QCM-D is a surface sensitive technique that measures changes in (dynamically coupled) mass and viscoelastic properties close to the sensor surface, within a distance into the cell that is usually only a fraction of its size. In this work, QCM-D was combined with light microscopy to study in situ cell attachment and spreading. Overtone-dependent changes of the QCM-D responses (frequency and dissipation shifts) were first recorded, as fibroblast cells attached to protein-coated sensors in a window equipped flow module. Then, as the cell layer had stabilised, morphological changes were induced in the cells by injecting cytochalasin D. This caused changes in the QCM-D signals that were reversible in the sense that they disappeared upon removal of cytochalasin D. These results are compared to other cell QCM-D studies. Our results stress the combination of QCM-D and light microscopy to help interpret QCM-D results obtained in cell assays and thus suggests a direction to develop the QCM-D technique as an even more useful tool for real-time cell studies.     

Productivity and nutritive value of Brachiaria decumbens and performance of dairy heifers in a long‐term silvopastoral system

Silvopastoral system (SPS) has been suggested to ensure sustainability in animal production systems in tropical ecosystems. The objective of this study was to evaluate the productive and nutritive value traits of Brachiaria decumbens and performance of dairy heifers in SPS and open pasture (OP) during the summer and autumn of two consecutive years in the SPS and OP (17th and 18th years after establishment). Experimental design was a randomized complete block with two treatments (SPS and OP) and three replications in subdivided plots with repeated measures in time. There was reduction in the tiller population density, total and green forage mass, and total and green forage bulk density in the SPS when compared to the OP in the summer. In the autumn, no difference was observed between the systems. Shading in SPS increased crude protein content of the grass pasture by 25% and 33% when compared to the OP during the first and second experimental years, respectively, and reduced neutral detergent fibre content. However, it did not change acid detergent fibre or lignin content. OP provided higher stocking rate and weight gain per area than the SPS. Average daily gain was higher in the OP in the second experimental year. Severe shading conditions should be avoided, because in the long ‐ term they may threaten the persistence and sustainability of pasture in the SPS. It is recommended to plant a low density of trees or implement management strategies of the tree component by either thinning or pruning over the years after planting.     

Lithothamnion muelleri Controls Inflammatory Responses,Target Organ Injury and Lethality Associated with Graft-versus-Host Disease in Mice

Lithothamnion muelleri (Hapalidiaceae) is a marine red alga, which is a member of a group of algae with anti-inflammatory, antitumor, and immunomodulatory properties. The present study evaluated the effects of treatment with Lithothamnion muelleri extract (LM) in a model of acute graft-versus-host disease (GVHD), using a model of adoptive splenocyte transfer from C57BL/6 donors into B6D2F1 recipient mice. Mice treated with LM showed reduced clinical signs of disease and mortality when compared with untreated mice. LM-treated mice had reduced tissue injury, less bacterial translocation, and decreased levels of proinflammatory cytokines and chemokines (interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), chemokine (C-C motif) ligand 2 (CCL2), chemokine (C-C motif) ligand 3 (CCL3) and chemokine (C-C motif) ligand 5 (CCL5)). The polysaccharide-rich fraction derived from LM could inhibit leukocyte rolling and adhesion in intestinal venules, as assessed by intravital microscopy. LM treatment did not impair the beneficial effects of graft-versus-leukaemia (GVL). Altogether, our studies suggest that treatment with Lithothamnion muelleri has a potential therapeutic application in GVHD treatment.     

Molecular Analysis of Phytophthora infestans (Mont.) de Bary from Chapingo,Mexico. Phylogeographic Referential

An Andean origin of Phytophthora infestans was originally based on written records, and recently inferred from mitochondrial and nuclear gene genealogies, which emphasize a common South American origin of the European isolates, and absence of the I-b haplotype in Mexico (recently identified in Tlaxcala, Mexico). In order to compare such results, we sequenced the nuclear genes β-tubulin, Ras, and Intron Ras, and rDNA ITS regions in 24 P. infestans genotypes isolated, identified, and selected, during three crop growing cycles in Chapingo, Mexico. The pathogen showed homology and 100 % sequence similarities to the ones reported for the Andes and Europe β-tubulin, Ras, and Intron Ras genes. Results for the ITS region were inconclusive. The molecular oomycete similarities from the three geographical regions (South America, Europe, and Mexico), and the Mexican genotypes characteristics reported in the species supports the hypothesis of the location of P. infestans diversity and a center of dispersal in the central highlands of Mexico.     

The V-antigen of Yersinia forms a distinct structure at the tip of injectisome needles

Many pathogenic bacteria use injectisomes to deliver effector proteins into host cells through type III secretion. Injectisomes consist of a basal body embedded in the bacterial membranes and a needle. In Yersinia, translocation of effectors requires the YopB and YopD proteins, which form a pore in the target cell membrane, and the LcrV protein, which assists the assembly of the pore. Here we report that LcrV forms a distinct structure at the tip of the needle, the tip complex. This unique localization of LcrV may explain its crucial role in the translocation process and its efficacy as the main protective antigen against plague.     

Mycobacterial Ku and ligase proteins constitute a two-component NHEJ repair machine

In mammalian cells, repair of DNA double-strand breaks (DSBs) by nonhomologous end-joining (NHEJ) is critical for genome stability. Although the end-bridging and ligation steps of NHEJ have been reconstituted in vitro, little is known about the end-processing reactions that occur before ligation. Recently, functionally homologous end-bridging and ligation activities have been identified in prokarya. Consistent with its homology to polymerases and nucleases, we demonstrate that DNA ligase D from Mycobacterium tuberculosis (Mt-Lig) possesses a unique variety of nucleotidyl transferase activities, including gap-filling polymerase, terminal transferase, and primase, and is also a 3' to 5' exonuclease. These enzyme activities allow the Mt-Ku and Mt-Lig proteins to join incompatible DSB ends in vitro, as well as to reconstitute NHEJ in vivo in yeast. These results demonstrate that prokaryotic Ku and ligase form a bona fide NHEJ system that encodes all the recognition, processing, and ligation activities required for DSB repair.     

Signal-driven computations in speech processing

Learning a language requires both statistical computations to identify words in speech and algebraic-like computations to discover higher level (grammatical) structure. Here we show that these computations can be influenced by subtle cues in the speech signal. After a short familiarization to a continuous speech stream, adult listeners are able to segment it using powerful statistics, but they fail to extract the structural regularities included in the stream even when the familiarization is greatly extended. With the introduction of subliminal segmentation cues, however, these regularities can be rapidly captured.     

Identification of a DNA nonhomologous end-joining complex in bacteria

In eukaryotic cells, double-strand breaks (DSBs) in DNA are generally repaired by the pathway of homologous recombination or by DNA nonhomologous end joining (NHEJ). Both pathways have been highly conserved throughout eukaryotic evolution, but no equivalent NHEJ system has been identified in prokaryotes. The NHEJ pathway requires a DNA end-binding component called Ku. We have identified bacterial Ku homologs and show that these proteins retain the biochemical characteristics of the eukaryotic Ku heterodimer. Furthermore, we show that bacterial Ku specifically recruits DNA ligase to DNA ends and stimulates DNA ligation. Loss of these proteins leads to hypersensitivity to ionizing radiation in Bacillus subtilis. These data provide evidence that many bacteria possess a DNA DSB repair apparatus that shares many features with the NHEJ system of eukarya and suggest that this DNA repair pathway arose before the prokaryotic and eukaryotic lineages diverged.