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In this work, LIBS (Laser Induced Breakdown Spectroscopy) capability to operate in stand-off configuration, without the need of a direct contact with the sample, has been coupled with the calibration-free (CF) approach to LIBS data analysis. The latter does not require the use of standard calibration. The feasibility of the calibration free method on stand-off LIBS spectra has been thereby tested. The quantitative analysis was performed on samples of two well-known meteorites, Toluca (iron meteorite) and Sahara 98222 (L6 chondrite) by using a stand-off configuration at a distance of 5 m. The plasma temperature and the electron number densities were determined for each sample and for each laser shot in order to apply the CF method. For the Toluca meteorite sample Fe, Ni and Co content was quantified during the depth profile. For the Sahara 98222 major and minor elements (Fe, Mg, Si, Na, Ti, Al, Cr, Mn, Ca, Ni, Co) were analysed by averaging different meteorite zones because of its strong inhomogeneity. Results demonstrate the possibility of remote analysis of minor bodies and space debris. [ABSTRACT FROM AUTHOR]

There is a growing interest in the development of sensors use in exploration of the deep ocean. Techniques for the chemical analysis of submerged solids are of special interest, as they show promise for subsea mining applications where a rapid sorting of materials found in the sea bottom would improve efficiency. Laser-Induced Breakdown Spectroscopy (LIBS) has demonstrated potential for this application thanks to its unique capability of providing the atomic composition of submerged solids. Here we present a study on the parameters that affect the spectral response of metallic targets in an oceanic pressure environment. Following laser excitation of the solid, the plasma persistence and the cavitation bubble size are considerably reduced as the hydrostatic pressure increases. These effects are of particular concern in dual pulse excitation as reported here, where a careful choice of the interpulse timing is required. Shadowgraphic images of the plasma demonstrate that cavitation bubbles are formed early after the plasma onset and that the effect of hydrostatic pressure is negligible during the early stage of plasma expansion. Contrarily to what is observed at atmospheric pressure, emission spectra observed at high pressures are characterized by self-absorbed atomic lines on continuum radiation resulting from strong radiative recombination in the electron-rich confined environment. This effect is much less evident with ionic lines due to the much higher energy of the levels involved and ionization energy of ions, as well as to the lower extent of absorption effects occurring in the inner part of the plasma, where ionized species are more abundant. As a result of the smaller shorter-lived cavitation bubble, the LIBS intensity enhancement resulting from dual pulse excitation is reduced when the applied pressure increases. [ABSTRACT FROM AUTHOR]

Solubility, density and the refractive index were determined for saturated solutions of the systems CuSO 4 + H 2 SO 4 + (2-propanol + H 2 O), CuSO 4 + H 2 SO 4 + (FeSO 4 + H 2 O) and CuSO 4 + CuCl 2 + (H 2 SO 4 + H 2 O). The experimental study was carried out at 298.15 K, and for different mass ratios of 0.04 of 2-propanol/water, 0.02 and 0.002 of ferrous sulfate/water and 0.1 of sulfuric acid/water, respectively. 2-Propanol has a drowning-out effect, decreasing copper sulphate solubility. No remarkable effect was found when adding FeSO 4 . With the experimental results obtained, the phase diagrams and their regions were determined. The most important precipitated salt for all the analyzed systems is copper sulfate pentahydrate. The experimental data on solubility and physical properties of the copper sulphate as a function of the sulfuric acid concentration were correlated with empirical equations for all the systems. [ABSTRACT FROM AUTHOR]

Thelow availability of fresh water in Chilean deserts has encouragedresearch into new alternative water sources in the mining industry.One of alternative is the use of saline water; however, there arefew data about this new approach. For the production of copper sulfatepentahydrated (CuSO4·5H2O) crystals, theeffect of NaCl in the crystallization area must be known; therefore,the solid–liquid equilibrium of the CuSO4+ NaCl+ (H2O or H2SO4/H2O) systemat 298.15 K was experimentally determined by the wet residue method.The density and refractive index of saturated solutions were alsomeasured. The addition of NaCl has a clear effect on the solid–liquidequilibrium, promoting the formation of solid phases. In the phaseequilibrium diagram that includes sulfuric acid, the pH affects thesolubility curve. Using the information of the new phase diagrams,six simulation cases, varying in NaCl content and the addition ofsulfuric acid in the feed, were evaluated in terms of mass and energybalance. The addition of sulfuric acid increases the total fluxesbut decreases the heat requirement. Contrary to the expected results,the yields for the cases that include sulfuric acid were lower. Thebest results were obtained using a 50% NaCl pulp, which produced thelowest total flux and a high yield of 75.6%. [ABSTRACT FROM AUTHOR]

Abstract: Plastic changes in the adult mammal hippocampus can be altered by many factors and perhaps the most well-documented is stress. Stress and elevated corticosterone levels have been shown to decrease hippocampal neurogenesis and decrease the complexity of CA3 pyramidal neurons. However, the extent of these changes in relation to low and moderately elevated levels of corticosterone has yet to be fully investigated. Therefore, the aim of the present study was to determine how low to moderately elevated circulating corticosterone levels affect dendritic morphology of CA3 pyramidal cells and hippocampal neurogenesis in adult male rats. To do this, three groups of adult male Wistar rats were used: (1) Sham-operated, (2) Adrenalectomized (ADX), and (3) ADX+corticosterone replacement. Primary results show that adrenalectomy, but not moderately elevated levels of corticosterone replacement, resulted in significant atrophy of CA3 pyramidal neurons. Interestingly, moderate corticosterone replacement resulted in significantly more surviving new cells in the dentate gyrus when compared to sham controls. This work shows that circulating levels of corticosterone differentially affect plasticity in the CA3 region and the dentate gyrus. [Copyright &y& Elsevier]

In higher plants, ammonium is assimilated into amino acids through the glutamine synthetase (GS)/glutamate synthase (GOGAT) cycle. This metabolic cycle is distributed in different cellular compartments in conifer seedlings: glutamine synthesis occurs in the cytosol and glutamate synthesis within the chloroplast. A method for preparing intact chloroplasts of pine cotyledons is presented with the aim of identifying a glutamine–glutamate translocator. Glutamine–glutamate exchange has been studied using the double silicone layer system, suggesting the existence of a translocator that imports glutamine into the chloroplast and exports glutamate to the cytoplasm. The translocator identified is specific for glutamine and glutamate, and the kinetic constants for both substrates indicate that it is unsaturated at intracellular concentrations. Thus, the experimental evidence obtained supports the model of the GS/GOGAT cycle in developing pine seedlings that accounts for the stoichiometric balance of metabolites. As a result, the efficient assimilation of free ammonia produced by photorespiration, nitrate reduction, storage protein mobilisation, phenylpropanoid pathway or S-adenosylmethionine synthesis is guaranteed. [ABSTRACT FROM AUTHOR]

Ripening is a complex physiological process that involves changes in reactive nitrogen and oxygen species that govern the shelf-life and quality of fruits. Nitric oxide (NO)-dependent changes in the sweet pepper fruit transcriptome were determined by treating fruits at the initial breaking point stage with NO gas. Fruits were also harvested at the immature (green) and ripe (red) stages. Fruit ripening in the absence of NO resulted in changes in the abundance of 8805 transcripts whose function could be identified. Among these, functional clusters associated with reactive oxygen/nitrogen species and lipid metabolism were significantly modified. NO treatment resulted in the differential expression of 498 genes framed within these functional categories. Biochemical analysis revealed that NO treatment resulted in changes in fatty acid profiling, glutathione and proline contents, and the extent of lipid peroxidation, as well as increases in the activity of ascorbate peroxidase and lipoxygenase. These data provide supporting evidence for the crucial role of NO in the ripening of pepper fruit. [ABSTRACT FROM AUTHOR]

The excessive accumulation of chloride (Cl−) in leaves due to salinity is frequently related to decreased yield in citrus. Two salt tolerance experiments to detect quantitative trait loci (QTLs) for leaf concentrations of Cl−, Na+, and other traits using the same reference progeny derived from the salt-tolerant Cleopatra mandarin (Citrus reshni) and the disease-resistant donor Poncirus trifoliata were performed with the aim to identify repeatable QTLs that regulate leaf Cl− (and/or Na+) exclusion across independent experiments in citrus, as well as potential candidate genes involved. A repeatable QTL controlling leaf Cl− was detected in chromosome 6 (LCl-6), where 23 potential candidate genes coding for transporters were identified using the C. clementina genome as reference. Transcriptomic analysis revealed two important candidate genes coding for a member of the nitrate transporter 1/peptide transporter family (NPF5.9) and a major facilitator superfamily (MFS) protein. Cell wall biosynthesis- and secondary metabolism-related processes appeared to play a significant role in differential gene expression in LCl-6. Six likely gene candidates were mapped in LCl-6, showing conserved synteny in C. reshni. In conclusion, markers to select beneficial Cleopatra mandarin alleles of likely candidate genes in LCl-6 to improve salt tolerance in citrus rootstock breeding programs are provided. [ABSTRACT FROM AUTHOR]

Background: Gene expression analyses demand appropriate reference genes (RGs) for normalization, in order to obtain reliable assessments. Ideally, RG expression levels should remain constant in all cells, tissues or experimental conditions under study. Housekeeping genes traditionally fulfilled this requirement, but they have been reported to be less invariant than expected; therefore, RGs should be tested and validated for every particular situation. Microarray data have been used to propose new RGs, but only a limited set of model species and conditions are available; on the contrary, RNA-seq experiments are more and more frequent and constitute a new source of candidate RGs.Results: An automated workflow based on mapped NGS reads has been constructed to obtain highly and invariantly expressed RGs based on a normalized expression in reads per mapped million and the coefficient of variation. This workflow has been tested with Roche/454 reads from reproductive tissues of olive tree (Olea europaea L.), as well as with Illumina paired-end reads from two different accessions of Arabidopsis thaliana and three different human cancers (prostate, small-cell cancer lung and lung adenocarcinoma). Candidate RGs have been proposed for each species and many of them have been previously reported as RGs in literature. Experimental validation of significant RGs in olive tree is provided to support the algorithm.Conclusion: Regardless sequencing technology, number of replicates, and library sizes, when RNA-seq experiments are designed and performed, the same datasets can be analyzed with our workflow to extract suitable RGs for subsequent PCR validation. Moreover, different subset of experimental conditions can provide different suitable RGs. [ABSTRACT FROM AUTHOR]

Many harvested timber species of tropical forests have not been regenerating sufficiently and have thus made post-harvesting silvicultural treatments necessary. This study analysed the profitability of sawnwood produced through four treatments in seedlings and saplings naturally present or planted in logging gaps. The treatments were (1) standard procedures of reduced-impact logging or control, (2) tending of the naturally established regeneration, (3) enrichment planting 1 (EP-1) and (4) enrichment planting 2 (EP-2). In EP-1 seedlings were planted in 2-year-old gaps keeping logging residuals inside the gap while in EP-2 seedlings were planted in 1-year-old gaps with residuals removed. Species of EP-2 had higher financial value and growth rates than species in EP-1. Growth rates of treated individuals were projected in 30 and 60 years to simulate sawnwood production. With increases of 25 and 50% in growth rates, 500% in timber prices, and interest rates of 4 and 6% year-1, tending and enrichment planting could profit at year 60. These silvicultural treatments, with technological improvements, can become even more financially profitable for forest managers and investors. [ABSTRACT FROM AUTHOR]

Physical and genetic maps of chickpea a QTL related to Ascochyta blight resistance and located in LG2 (QTL) have been constructed. Single-copy markers based on candidate genes located in the Ca2 pseudomolecule were for the first time obtained and found to be useful for refining the QTL position. The location of the QTL peak was linked to an ethylene insensitive 3-like gene ( Ein3). The Ein3 gene explained the highest percentage of the total phenotypic variation for resistance to blight (44.3 %) with a confidence interval of 16.3 cM. This genomic region was predicted to be at the Ca2 physical position 32-33 Mb, comprising 42 genes. Candidate genes located in this region include Ein3, Avr9/Cf9 and Argonaute 4, directly involved in disease resistance mechanisms. However, there are other genes outside the confidence interval that may play a role in the blight resistance pathway. The information reported in this paper will facilitate the development of functional markers to be used in the screening of germplasm collections or breeding materials, improving the efficiency and effectiveness of conventional breeding methods. [ABSTRACT FROM AUTHOR]

Background: Nowadays, microarray gene expression analysis is a widely used technology that scientists handle but whose final interpretation usually requires the participation of a specialist. The need for this participation is due to the requirement of some background in statistics that most users lack or have a very vague notion of. Moreover, programming skills could also be essential to analyse these data. An interactive, easy to use application seems therefore necessary to help researchers to extract full information from data and analyse them in a simple, powerful and confident way. Results: PreP+07 is a standalone Windows XP application that presents a friendly interface for spot filtration, inter- and intra-slide normalization, duplicate resolution, dye-swapping, error removal and statistical analyses. Additionally, it contains two unique implementation of the procedures -- double scan and Supervised Lowess-, a complete set of graphical representations -- MA plot, RG plot, QQ plot, PP plot, PN plot -- and can deal with many data formats, such as tabulated text, GenePix GPR and ArrayPRO. PreP+07 performance has been compared with the equivalent functions in Bioconductor using a tomato chip with 13056 spots. The number of differentially expressed genes considering p-values coming from the PreP+07 and Bioconductor Limma packages were statistically identical when the data set was only normalized; however, a slight variability was appreciated when the data was both normalized and scaled. Conclusion: PreP+07 implementation provides a high degree of freedom in selecting and organizing a small set of widely used data processing protocols, and can handle many data formats. Its reliability has been proven so that a laboratory researcher can afford a statistical pre-processing of his/her microarray results and obtain a list of differentially expressed genes using PreP+07 without any programming skills. All of this gives support to scientists that have been using previous PreP releases since its first version in 2003. [ABSTRACT FROM AUTHOR]

The photosynthetic protist Euglena gracilis contains chloroplasts surrounded by three membranes which arise from secondary endosymbiosis. The genes petA and petD, encoding cytochrome f and subunit IV of the cytochrome bf complex, normally present in chloroplast genomes, are lacking from the chloroplast DNA (cpDNA) of E. gracilis. The bf complex of E. gracilis was isolated, and the identities of cytochrome f and subunit IV were established immunochemically, by heme-specific staining, and by Edman degradation. Based on N-terminal and conserved internal protein sequences, primers were designed and used for PCR gene amplification and cDNA sequencing. The complete sequence of the petA cDNA and the partial sequence of the petD cDNA from E. gracilis are described. Evidence is provided that in this protist, the petA and petD genes have migrated from the chloroplast to the nucleus. Both genes exhibit a typical nuclear codon usage, clearly distinct from the usage of chloroplast genes. The petA gene encodes an atypical cytochrome f, with a unique insertion of 62 residues not present in other f-type cytochromes. The petA gene also acquired a region that encodes a large tripartite chloroplast transit peptide (CTP), which is thought to allow the import of apocytochrome f through the three-membrane envelope of E. gracilis chloroplasts. This is the first description of petA and petD genes that are nucleus-localized. [Copyright &y& Elsevier]

The sequence of 12 SCAR markers obtained for olive-tree was analysed in the seek of information about the nature of the RAPD polymorphisms in olive-tree. The absence of internal repetitions of the RAPD primer corroborates that the polymorphisms were not obtained by alternative priming nor by rearrangements. We have found a high degree repetition within each sequence which is a reflect of the repetitive nature of the RAPD polymorphisms. Most the sequence corresponded to unknown DNA sequences, although some of the SCAR markers like GH1, PH2 or PF84 present clear homology or identity to other repetitive sequences (mainly microsatellites) from other plants or animals. This suggests that repetitive sequences could be more conserved among species than initially thought. As expected, none of the markers presented any open reading frame, although PJ65 revealed a significant homology to the gene and pseudogene of a lectin receptor in Arabidopsis. Taken together, the results described in this paper demonstrate that the polymorphisms detected by SCAR/RAPD methods are mainly repetitive in olive-tree, and that most of these repetitions corresponded to microsatellite DNA. [ABSTRACT FROM AUTHOR]

Objective To determine persistence and variability of colonization with Pseudomonas aeruginosa in cystic fibrosis patients over long time periods, and to look for possible cross-colonization. Methods In total, 469 Pseudomonas aeruginosa isolates were obtained from 30 patients during the period from April 1994 to April 1996. The sources were mainly sputum and a few deep throat swabs. All grown strains dissimilar in macromorphology were processed separately. Typing with PFGE was carried out by contour-clamped homogeneous electric field electrophoresis. Genomic DNA was subjected to the rare-cutting restriction enzyme SpeI. For pyocin typing, the procedure described by Fyfe was applied. Results After typing with PFGE, we observed 40 restriction profiles. Eighteen different pyocin types were found. The most frequent pyocin type was type 3, followed by types 1 and 5. Twenty-two patients were persistently colonized by one clone specific and different for each patient, and four were co-colonized by a second clone also different for each of these patients. Cross-colonization had apparently been rare in the cystic fibrosis center of Leipzig. Conclusions Typing with PFGE is well suited for detailed investigations of colonization with Pseudomonas aeruginosa in cystic fibrosis patients. Pyocin typing can provide additional information for epidemiologic purposes. [ABSTRACT FROM AUTHOR]

The synthetic auxin 2,4‐dichlorophenoxyacetic acid (2,4‐D) functions as an agronomic weed control herbicide. High concentrations of 2,4‐D induce plant growth defects, particularly leaf epinasty and stem curvature. Although the 2,4‐D triggered reactive oxygen species (ROS) production, little is known about its signalling. In this study, by using a null mutant in peroxisomal acyl CoA oxidase 1 (acx1‐2), we identified acyl‐coenzyme A oxidase 1 (ACX1) as one of the main sources of ROS production and, in part, also causing the epinastic phenotype following 2,4‐D application. Transcriptomic analyses of wild type (WT) plants after treatment with 2,4‐D revealed a ROS‐related peroxisomal footprint in early plant responses, while other organelles, such as mitochondria and chloroplasts, are involved in later responses. Interestingly, a group of 2,4‐D‐responsive ACX1‐dependent transcripts previously associated with epinasty is related to auxin biosynthesis, metabolism, and signalling. We found that the auxin receptor auxin signalling F‐box 3 (AFB3), a component of Skp, Cullin, F‐box containing complex (SCF) (ASK‐cullin‐F‐box) E3 ubiquitin ligase complexes, which mediates auxin/indole acetic acid (AUX/IAA) degradation by the 26S proteasome, acts downstream of ACX1 and is involved in the epinastic phenotype induced by 2,4‐D. We also found that protein degradation associated with ubiquitin E3‐RING and E3‐SCF‐FBOX in ACX1‐dependent signalling in plant responses to 2,4‐D is significantly regulated over longer treatment periods. Summary Statement: 2,4‐D induced a reactive oxygen species (ROS)‐related peroxisomal footprint in early plant responses. ACX1 is one of the main sources of ROS production and associated epinasty phenotype following 2,4‐D application in plants. ACX1‐dependent signalling in plant responses to 2,4‐D point to protein degradation associated with auxin signalling at the early response and with ubiquitin E3‐RING and E3‐SCF‐FBOX/proteasome as key processes in the development of epinasty over longer treatment periods. [ABSTRACT FROM AUTHOR]

The integration of physical and high-density genetic maps is a very useful approach to achieve chromosome-level genome assemblies. Here, the genome of a male Senegalese sole (Solea senegalensis) was de novo assembled and the contigs were anchored to a high-quality genetic map for chromosome-level scaffolding. Hybrid assembled genome was 609.3 Mb long and contained 3403 contigs with a N50 of 513 kb. The linkage map was constructed using 16,287 informative SNPs derived from ddRAD sequencing in 327 sole individuals from five families. Markers were assigned to 21 linkage groups with an average number of 21.9 markers per megabase. The anchoring of the physical to the genetic map positioned 1563 contigs into 21 pseudo-chromosomes covering 548.6 Mb. Comparison of genetic and physical distances indicated that the average genome-wide recombination rate was 0.23 cM/Mb and the female-to-male ratio 1.49 (female map length: 2,698.4 cM, male: 2,036.6 cM). Genomic recombination landscapes were different between sexes with crossovers mainly concentrated toward the telomeres in males while they were more uniformly distributed in females. A GWAS analysis using seven families identified 30 significant sex-associated SNP markers located in linkage group 18. The follicle-stimulating hormone receptor appeared as the most promising locus associated with sex within a region with very low recombination rates. An incomplete penetrance of sex markers with males as the heterogametic sex was determined. An interspecific comparison with other Pleuronectiformes genomes identified a high sequence similarity between homologous chromosomes, and several chromosomal rearrangements including a lineage-specific Robertsonian fusion in S. senegalensis. [ABSTRACT FROM AUTHOR]

The Senegalese sole (Solea senegalensis) is an economically important flatfish species. In this study, a genome draft was analyzed to identify microsatellite (SSR) markers for whole-genome genotyping. A subset of 224 contigs containing SSRs were preselected and validated by using a de novo female hybrid assembly. Overall, the SSR density in the genome was 886.7 markers per megabase of genomic sequences and the dinucleotide motif was the most abundant (52.4%). In silico comparison identified a set of 108 SSRs (with di-, tetra- or pentanucleotide motifs) widely distributed in the genome and suitable for primer design. A total of 106 markers were structured in thirteen multiplex PCR assays (with up to 10-plex) and the amplification conditions were optimized with a high-quality score. Main genetic diversity statistics and genotyping reliability were assessed. A subset of 40 high polymorphic markers were selected to optimize four supermultiplex PCRs (with up to 11-plex) for pedigree analysis. Theoretical exclusion probabilities and real parentage allocation tests using parent–offspring information confirmed their robustness and effectiveness for parental assignment. These new SSR markers were combined with previously published SSRs (in total 229 makers) to construct a new and improved integrated genetic map containing 21 linkage groups that matched with the expected number of chromosomes. Synteny analysis with respect to C. semilaevis provided new clues on chromosome evolution in flatfish and the formation of metacentric and submetacentric chromosomes in Senegalese sole. [ABSTRACT FROM AUTHOR]

Body shape is a complex and plastic trait with a high impact on fish performance and commercialization. Shape is particularly relevant in flatfish with highly asymmetric bodies. In this study we investigated the skeleton features, transcriptomic profiles and genomic regions in two Senegalese sole groups from a full-sib family with positive (named as HE) or negative (LE) breeding values for body ellipticity (ELL). Soles from HE group were heavier and displayed more elongated bodies than LE. A skeleton analysis showed that HE had a higher number of vertebrae, longer vertebral bodies and intervertebral spaces and a lower incidence of skeletal fusions than LE. With respect to the gender effect, females showed a higher height of ventral body and some differences in the ratio urostyle to skeleton length than males for similar ELL values. The PCA analysis using 28 dimensionless morphological traits indicated that PC1 and PC2 explained 51.3% of the total variance and they were mainly associated with ELL and gender variation, respectively. Expression analysis using two factors (ELL and gender) revealed a major effect of gender in muscle, pterygiophores and liver with 6764, 6506 and 7559 differentially expressed transcripts (DETs) between sexes, respectively. In brain, only 16 DETs were identified. When HE and LE were compared controlling by gender, we identified the subset of DETs with conserved responses in both sexes or those regulated in a sex-specific manner in each tissue. A comparison across tissues identified 31 ubiquitous transcripts as differentially expressed in at least two of them. Functional analysis showed that liver and pterygiophores had the highest number of enriched gene ontology categories (24 and 14, respectively). Some of these enriched pathways were related to regulation of cell cycle, cell structure and shape, locomotory behavior, immune system, hormonal responses and homeostasis. A BSR-Seq analysis confirmed five significant genomic windows associated with ELL by using the tricube-smoothed G' and Fst statistics. Windows located in linkage groups (LG) 5 and LG14 included the malt1 and vtg3 genes, respectively, that were also detected within the subset of ubiquitous DETs. Moreover, malt1 appeared located very close to the bmpr1b gene within the significance hotspot of LG5 indicating that mediators of the inflammatory responses and BMP signaling pathway could be responsible for the differences found in the skeleton features and body ellipticity. • Body ellipticity was associated with changes in skeleton features. • Transcriptional responses related to shape plasticity were investigated in four tissues. • Gender highly modulated gene expression defining sex-specific responses to ellipticity. • Five significant genomic windows that explained body ellipticity were identified. • malt1 , bmpr1b and vtg3 were proposed as main gene candidates to regulate ellipticity. [ABSTRACT FROM AUTHOR]

Background: Podosphaera xanthii is the main causal agent of powdery mildew disease in cucurbits and is responsible for important yield losses in these crops worldwide. Powdery mildew fungi are obligate biotrophs. In these parasites, biotrophy is determined by the presence of haustoria, which are specialized structures of parasitism developed by these fungi for the acquisition of nutrients and the delivery of effectors. Detailed molecular studies of powdery mildew haustoria are scarce due mainly to difficulties in their isolation. Therefore, their analysis is considered an important challenge for powdery mildew research. The aim of this work was to gain insights into powdery mildew biology by analysing the haustorial transcriptome of P. xanthii. Results: Prior to RNA isolation and massive-scale mRNA sequencing, a flow cytometric approach was developed to isolate P. xanthii haustoria free of visible contaminants. Next, several commercial kits were used to isolate total RNA and to construct the cDNA and Illumina libraries that were finally sequenced by the Illumina NextSeq system. Using this approach, the maximum amount of information from low-quality RNA that could be obtained was used to accomplish the de novo assembly of the P. xanthii haustorial transcriptome. The subsequent analysis of this transcriptome and comparison with the epiphytic transcriptome allowed us to identify the importance of several biological processes for haustorial cells such as protection against reactive oxygen species, the acquisition of different nutrients and genetic regulation mediated by non-coding RNAs. In addition, we could also identify several secreted proteins expressed exclusively in haustoria such as cell adhesion proteins that have not been related to powdery mildew biology to date. Conclusions: This work provides a novel approach to study the molecular aspects of powdery mildew haustoria. In addition, the results of this study have also allowed us to identify certain previously unknown processes and proteins involved in the biology of powdery mildews that could be essential for their biotrophy and pathogenesis. [ABSTRACT FROM AUTHOR]

Humans have been modifying ecosystems since before the Holocene began ca. 12,000 years ago, even in Neotropical regions. The Amazon was once thought to be 'pristine' and only lightly impacted by Indigenous people before European colonization in the Americas (e.g., pre-Columbian); however, multiple lines of evidence have shown that Indigenous human activities over the past millennia have left ecological legacies on modern ecosystems. We review the various lines of evidence used to reconstruct pre-Columbian Indigenous human activity in Amazonia, and assess the spatial and temporal resolution and limits of each one of them. We suggest that a multi-proxy approach is always preferred, and that lines of evidence that cover overlapping yet discrete spatial and temporal scales can provide a robust and comprehensive assessment of the nuances of pre-Columbian Indigenous human activities in Amazonia, and how they affect modern ecosystems. • Evidence of pre-Columbian Indigenous human activity is more robust using a multi-disciplinary approach. • Pre-Columbian Indigenous human activity in Amazonia is reconstructed using a variety of data sources and proxies. • Each line of evidence has its own potential spatial and temporal resolution and spatial and temporal limit. [ABSTRACT FROM AUTHOR]

Pepper ( Capsicum annuum , L.) fruit ripening is characterized by important phenotypic and metabolic changes, with emission of volatile compounds, cell wall softening, chlorophyll degradation, and new pigments and proteins’ synthesis. Recent data have shown that reactive oxygen and nitrogen species (ROS/RNS) are involved in fruit ripening by regulating both gene expression and protein functions through post-translational modifications. In this work, using immature (green) and ripe (red) pepper fruits, RNA-sequencing was performed through an Illumina platform. Raw reads were pre-processed and assembled resulting in a representative transcriptome of 63,353 tentative transcripts (TTs), of which 29,716 had a unique orthologue. Differential expression analysis between green and red pepper fruits showed that 4,325 and 3,269 transcripts were down-and up-regulated respectively at ripening. Moreover, isobaric tags for relative and absolute quantification (iTRAQ®) was used as a proteomic approach, identifying 2,574 proteins, of which 1,189 showed differential expression. In this study, we have combined transcriptomic and proteomic data focusing on oxidative metabolism and antioxidative systems to gain insight into their role on fruit ripening. [ABSTRACT FROM AUTHOR]

Background: The advances in high-throughput sequencing technologies are allowing more and more de novo assembling of transcriptomes from many new organisms. Some degree of automation and evaluation is required to warrant reproducibility, repetitivity and the selection of the best possible transcriptome. Workflows and pipelines are becoming an absolute requirement for such a purpose, but the issue of assembling evaluation for de novo transcriptomes in organisms lacking a sequenced genome remains unsolved. An automated, reproducible and flexible framework called TransFlow to accomplish this task is described. Results: TransFlow with its five independent modules was designed to build different workflows depending on the nature of the original reads. This architecture enables different combinations of Illumina and Roche/454 sequencing data, and can be extended to other sequencing platforms. Its capabilities are illustrated with the selection of reliable plant reference transcriptomes and the assembling six transcriptomes (three case studies for grapevine leaves, olive tree pollen, and chestnut stem, and other three for haustorium, epiphytic structures and their combination for the phytopathogenic fungus Podosphaera xanthii). Arabidopsis and poplar transcriptomes revealed to be the best references. A common result regarding de novo assemblies is that Illumina paired-end reads of 100 nt in length assembled with OASES can provide reliable transcriptomes, while the contribution of longer reads is noticeable only when they complement a set of short, single-reads. Conclusions: TransFlow can handle up to 181 different assembling strategies. Evaluation based on principal component analyses allows its self-adaptation to different sets of reads to provide a suitable transcriptome for each combination of reads and assemblers. As a result, each case study has its own behaviour, prioritises evaluation parameters, and gives an objective and automated way for detecting the best transcriptome within a pool of them. Sequencing data type and quantity (preferably several hundred millions of 2×100 nt or longer), assemblers (OASES for Illumina, MIRA4 and EULER-SR reconciled with CAP3 for Roche/454) and strategy (preferably scaffolding with OASES, and probably merging with Roche/454 when available) arise as the most impacting factors. [ABSTRACT FROM AUTHOR]

Photobacterium damselae subsp. piscicida ( Pdp ) is an intracellular fish pathogen that causes photobacteriosis, a disease proven deadly in farmed fish worldwide. This work focuses on the analysis of genome sequences, chromosomes structure and gene contents of two strains from Sparus aurata (DI21) and Solea senegalensis (L091106-03H), isolated on the Spanish Atlantic coast. The comparative genomic analysis revealed that DI21 and L091106-03H share 98% of their genomes, including two virulence plasmids: pPHDP70 encoding siderophore piscibactin synthesis and pPHDP10 encoding the apoptotic toxin AIP56. Both genomes harbour a surprisingly large number of IS elements accounting for 12–17% of the total genome, representing an IS density of 0.15 elements per kb, one of the highest IS density values in a bacterial pathogen. This massive proliferation of ISs is responsible for the generation of a high number of pseudogenes that caused extensive loss of biological functions. Pseudogene formation is one of the main features of Pdp genome that explains most of the ecological and phenotypic differences with respect to its sibling subspecies P. damselae subsp. damselae and to other Vibrionaceae . Evidence was also found proving the existence of two chromosomal configurations depending on the origin of the strains: an European and an Asian/American types of genome organisation, reinforcing the idea of the existence of two geographically-linked clonal lineages in Pdp . In short, our study suggests that the host-dependent lifestyle of Pdp allowed massive IS proliferation and gene decay processes, which are major evolutionary forces in the shaping of the Pdp genome. [ABSTRACT FROM AUTHOR]

Nitric oxide is recognized as a signaling molecule involved in a broad range of physiological processes in plants including sexual reproduction. NO has been detected in the pollen grain at high levels and regulates pollen tube growth. Previous studies demonstrated that NO as well as ROS are produced in the olive reproductive tissues in a stage- and tissue-specific manner. The aim of this study was to assess the production of NO throughout the germination of olive ( Olea europaea L.) pollen in vitro . The NO fluorescent probe DAF-2DA was used to image NO production in situ, which was correlated to pollen viability. Moreover, by means of a fluorimetric assay we showed that growing pollen tubes release NO. GSNO -a mobile reservoir of NO, formed by the S -nitrosylation of NO with reduced glutathione (GSH) - was for the first time detected and quantified at different stages of pollen tube growth using a LC-ES/MS analysis. Exogenous NO donors inhibited both pollen germination and pollen tube growth and these effects were partially reverted by the specific NO-scavenger c-PTIO. However, little is known about how NO affects the germination process. With the aim of elucidating the putative relevance of protein S -nitrosylation and Tyr-nitration as important post-translational modifications in the development and physiology of the olive pollen, a de novo assembled and annotated reproductive transcriptome from olive was challenged in silico for the putative capability of transcripts to become potentially modified by S -nitrosylation/Tyr-nitration according to well-established criteria. Numerous gene products with these characteristics were identified, and a broad discussion as regards to their potential role in plant reproduction was built after their functional classification. Moreover, the importance of both S -nitrosylation/Tyr-nitrations was experimentally assessed and validated by using Western blotting, immunoprecipitation and proteomic approaches. [ABSTRACT FROM AUTHOR]

Population genetic studies in tropical plants are often challenging because of limited information on taxonomy, phylogenetic relationships and distribution ranges, scarce genomic information and logistic challenges in sampling. We describe a strategy to develop robust and widely applicable genetic markers based on a modest development of genomic resources in the ancient tropical tree species Symphonia globulifera L.f. (Clusiaceae), a keystone species in African and Neotropical rainforests. We provide the first low-coverage (11X) fragmented draft genome sequenced on an individual from Cameroon, covering 1.027 Gbp or 67.5% of the estimated genome size. Annotation of 565 scaffolds (7.57 Mbp) resulted in the prediction of 1046 putative genes (231 of them containing a complete open reading frame) and 1523 exact simple sequence repeats ( SSRs, microsatellites). Aligning a published transcriptome of a French Guiana population against this draft genome produced 923 high-quality single nucleotide polymorphisms. We also preselected genic SSRs in silico that were conserved and polymorphic across a wide geographical range, thus reducing marker development tests on rare DNA samples. Of 23 SSRs tested, 19 amplified and 18 were successfully genotyped in four S. globulifera populations from South America (Brazil and French Guiana) and Africa (Cameroon and São Tomé island, FST = 0.34). Most loci showed only population-specific deviations from Hardy-Weinberg proportions, pointing to local population effects (e.g. null alleles). The described genomic resources are valuable for evolutionary studies in Symphonia and for comparative studies in plants. The methods are especially interesting for widespread tropical or endangered taxa with limited DNA availability. [ABSTRACT FROM AUTHOR]

The extent to which pre-Columbian societies altered Amazonian landscapes is hotly debated. We performed a basin-wide analysis of pre-Columbian impacts on Amazonian forests by overlaying known archaeological sites in Amazonia with the distributions and abundances of 85 woody species domesticated by pre-Columbian peoples. Domesticated species are five times more likely than nondomesticated species to be hyperdominant. Across the basin, the relative abundance and richness of domesticated species increase in forests on and around archaeological sites. In southwestern and eastern Amazonia, distance to archaeological sites strongly influences the relative abundance and richness of domesticated species. Our analyses indicate that modern tree communities in Amazonia are structured to an important extent by a long history of plant domestication by Amazonian peoples. [ABSTRACT FROM AUTHOR]

Salmonella Typhimurium is a food-borne pathogen that causes salmonellosis. When in contact with the host, neutrophils are rapidly recruited to act as first line of defense. To better understand the pathogenesis of this infection, we used an in vitro model of neutrophil infection to perform dual RNA-sequencing (both host and pathogen). In addition, and given that many pathogens interfere with kinase‐mediated phosphorylation in host signaling, we performed a phosphoproteomic analysis. The immune response was overall diminished in infected neutrophils, mainly JAK/STAT and toll-like receptor signaling pathways. We found decreased expression of proinflammatory cytokine receptor genes and predicted downregulation of the mitogen-activated protein (MAPK) signaling pathway. Also, Salmonella infection inhibited interferons I and II signaling pathways by upregulation of SOCS3 and subsequent downregulation of STAT1 and STAT2. Additionally, phosphorylation of PSMC2 and PSMC4, proteasome regulatory proteins, was decreased in infected neutrophils. Cell viability and survival was increased by p53 signaling, cell cycle arrest and NFkB-proteasome pathways activation. Combined analysis of RNA-seq and phosphoproteomics also revealed inhibited vesicle transport mechanisms mediated by dynein/dynactin and exocyst complexes, involved in ER-to-Golgi transport and centripetal movement of lysosomes and endosomes. Among the overexpressed virulence genes from Salmonella we found potential effectors responsible of these dysregulations, such as spiC, sopD2, sifA or pipB2, all of them involved in intracellular replication. Our results suggest that Salmonella induces (through overexpression of virulence factors) transcriptional and phosphorylation changes that increases neutrophil survival and shuts down immune response to minimize host response, and impairing intracellular vesicle transport likely to keep nutrients for replication and Salmonella -containing vacuole formation and maintenance. [Display omitted] • S. Typhimurium infection diminishes the immune response elicited by neutrophils. • S. Typhimurium infection increases survival of infected neutrophils. • S. Typhimurium infection inhibits vesicle transport mechanisms in neutrophils. [ABSTRACT FROM AUTHOR]

Background: In the era of DNA throughput sequencing, assembling and understanding gymnosperm mega-genomes remains a challenge. Although drafts of three conifer genomes have recently been published, this number is too low to understand the full complexity of conifer genomes. Using techniques focused on specific genes, gene models can be established that can aid in the assembly of gene-rich regions, and this information can be used to compare genomes and understand functional evolution. Results: In this study, gene capture technology combined with BAC isolation and sequencing was used as an experimental approach to establish de novo gene structures without a reference genome. Probes were designed for 866 maritime pine transcripts to sequence genes captured from genomic DNA. The gene models were constructed using GeneAssembler, a new bioinformatic pipeline, which reconstructed over 82 % of the gene structures, and a high proportion (85 %) of the captured gene models contained sequences from the promoter regulatory region. In a parallel experiment, the P. pinaster BAC library was screened to isolate clones containing genes whose cDNA sequence were already available. BAC clones containing the asparagine synthetase, sucrose synthase and xyloglucan endotransglycosylase gene sequences were isolated and used in this study. The gene models derived from the gene capture approach were compared with the genomic sequences derived from the BAC clones. This combined approach is a particularly efficient way to capture the genomic structures of gene families with a small number of members. Conclusions: The experimental approach used in this study is a valuable combined technique to study genomic gene structures in species for which a reference genome is unavailable. It can be used to establish exon/intron boundaries in unknown gene structures, to reconstruct incomplete genes and to obtain promoter sequences that can be used for transcriptional studies. A bioinformatics algorithm (GeneAssembler) is also provided as a Ruby gem for this class of analyses. [ABSTRACT FROM AUTHOR]

Pregnancy is a time of marked neural, physiological and behavioral plasticity in the female and is often a time when women are more vulnerable to stress and stress-related diseases, such as depression and anxiety. Unfortunately the impact of stress during gestation on neurobiological processes of the mother has yet to be fully determined, particularly with regard to changes in the hippocampus; a brain area that plays an important role in stress-related diseases. The present study aimed to determine how stress early in pregnancy may affect hippocampal plasticity in the pregnant female and whether these effects differ from those in virgin females. For this purpose, adult age-matched pregnant and virgin female Sprague–Dawley rats were divided into two conditions: (1) Control and (2) Stress. Females in the stress condition were restrained during days 5–11 of gestation and at matched time-points in virgin females. All pregnant females received an injection of bromodeoxyuridine (BrdU) on day 1 of gestation and were sacrificed 21 days later. The same procedure was carried out at matched time points in virgin females. Results show that for number of Ki67-immunoreactive (ir) cells and doublecortin (DCX)-ir cells, there were significant interactions between reproductive state (pregnant/virgin) and stress exposure ( p = .05, p = .04, respectively) with control virgin and stressed pregnant females having more Ki67-ir cells than control pregnant females and more DCX-ir cells than stressed virgin females. Results also show that pregnant females had significantly greater glucocorticoid receptor (GR) density in the CA1, CA3 and granule cell layer compared to virgin females. In addition, there was a main effect of stress on GR density in the CA3 region, with stressed females having significantly lower GR density compared to control females ( p = .01). This work adds to our understanding of how stress and reproductive state affect plasticity in the female hippocampus. [ABSTRACT FROM AUTHOR]

Background: Senegalese sole (Solea senegalensis) and common sole (S. solea) are two economically and evolutionary important flatfish species both in fisheries and aquaculture. Although some genomic resources and tools were recently described in these species, further sequencing efforts are required to establish a complete transcriptome, and to identify new molecular markers. Moreover, the comparative analysis of transcriptomes will be useful to understand flatfish evolution. Results: A comprehensive characterization of the transcriptome for each species was carried out using a large set of Illumina data (more than 1,800 millions reads for each sole species) and 454 reads (more than 5 millions reads only in S. senegalensis), providing coverages ranging from 1,384x to 2,543x. After a de novo assembly, 45,063 and 38,402 different transcripts were obtained, comprising 18,738 and 22,683 full-length cDNAs in S. senegalensis and S. solea, respectively. A reference transcriptome with the longest unique transcripts and putative non-redundant new transcripts was established for each species. A subset of 11,953 reference transcripts was qualified as highly reliable orthologs (>97% identity) between both species. A small subset of putative species-specific, lineage-specific and flatfish-specific transcripts were also identified. Furthermore, transcriptome data permitted the identification of single nucleotide polymorphisms and simple-sequence repeats confirmed by FISH to be used in further genetic and expression studies. Moreover, evidences on the retention of crystallins crybb1, crybb1-like and crybb3 in the two species of soles are also presented. Transcriptome information was applied to the design of a microarray tool in S. senegalensis that was successfully tested and validated by qPCR. Finally, transcriptomic data were hosted and structured at SoleaDB. Conclusions: Transcriptomes and molecular markers identified in this study represent a valuable source for future genomic studies in these economically important species. Orthology analysis provided new clues regarding sole genome evolution indicating a divergent evolution of crystallins in flatfish. The design of a microarray and establishment of a reference transcriptome will be useful for large-scale gene expression studies. Moreover, the integration of transcriptomic data in the SoleaDB will facilitate the management of genomic information in these important species. [ABSTRACT FROM AUTHOR]

Abstract: Exposure to prenatal stress (PS) can predispose individuals to the development of psychopathology later in life. We examined the effects of unpredictable chronic mild stress (CMS) exposure during adolescence on a background of PS in male and female Sprague-Dawley rats. PS induced more anxiety-like behavior in the elevated zero maze in both sexes, an effect that was normalized by subsequent exposure to CMS. Moreover, PS was associated with increased depression-like behavior in the forced swim test in males only. Conversely, sucrose intake was increased in PS males, whilst being decreased in females when consecutively exposed to PS and CMS. Hypothalamo–pituitary–adrenal (HPA) axis reactivity was affected in males only, with higher stress-induced plasma corticosterone levels after PS. Markedly, CMS normalized the effects of PS on elevated zero maze behavior as well as basal and stress-induced plasma corticosterone secretion. At the neurochemical level, both PS and CMS induced various sex-specific alterations in serotonin (5-HT) and tryptophan hydroxylase 2 (TPH2) immunoreactivity in the dorsal raphe nucleus, hippocampus and prefrontal cortex with, in line with the behavioral observations, more profound effects in male offspring. In conclusion, these findings show that prenatal maternal stress in Sprague-Dawley rats induces various anxiety- and depression-related behavioral and neuroendocrine changes, as well as alterations in central 5-HT and TPH2 function, predominantly in male offspring. Moreover, CMS exposure partially normalized the effects of previous PS experience, suggesting that the outcome of developmental stress exposure largely depends on the environmental conditions later in life and vice versa. [Copyright &y& Elsevier]

Recurrent breast cancer occurring after the initial treatment is associated with poor outcome. A bimodal relapse pattern after surgery for primary tumor has been described with peaks of early and late recurrence occurring at about 2 and 5 years, respectively. Although several clinical and pathological features have been used to discriminate between low- and high-risk patients, the identification of molecular biomarkers with prognostic value remains an unmet need in the current management of breast cancer. Using microarray-based technology, we have performed a microRNA expression analysis in 71 primary breast tumors from patients that either remained disease-free at 5 years post-surgery (group A) or developed early (group B) or late (group C) recurrence. Unsupervised hierarchical clustering of microRNA expression data segregated tumors in two groups, mainly corresponding to patients with early recurrence and those with no recurrence. Microarray data analysis and RT-qPCR validation led to the identification of a set of 5 microRNAs (the 5-miRNA signature) differentially expressed between these two groups: miR-149, miR-10a, miR-20b, miR-30a-3p and miR-342-5p. All five microRNAs were down-regulated in tumors from patients with early recurrence. We show here that the 5-miRNA signature defines a high-risk group of patients with shorter relapse-free survival and has predictive value to discriminate non-relapsing versus early-relapsing patients (AUC = 0.993, p-value<0.05). Network analysis based on miRNA-target interactions curated by public databases suggests that down-regulation of the 5-miRNA signature in the subset of early-relapsing tumors would result in an overall increased proliferative and angiogenic capacity. In summary, we have identified a set of recurrence-related microRNAs with potential prognostic value to identify patients who will likely develop metastasis early after primary breast surgery. [ABSTRACT FROM AUTHOR]

Abstract: Titania biomorphic ceramics were manufactured from wood preforms by a two steps process, i.e. infiltration and then pyrolysis in an inert atmosphere to produce porous TiC ceramics, followed by infiltration and pyrolysis in an air atmosphere where TiC was oxidized to TiO2. This technology provides a cost effective and eco-friendly route to advanced ceramic materials. The properties of biomorphic ceramics depend decisively on the synthesis parameters, that were optimized using a multivariate methodology for the design of experiments. Four variables (infiltration time, titanium isopropoxide and acetic acid proportion, and number of infiltrations before and after pyrolysis) were considered as factors in the synthesis optimization process. Interactions between these factors and their optimal levels were investigated using a two level factorial design. For evaluating the yields, a new method by Fourier transform infrared spectrometry (FTIR) has been developed for the direct determination of TiO2 by absorbance measurements in KBr pellets. The procedure is based on the use of the ratio between the absorbance of the characteristic band of titania and those of a nitrate internal standard added to samples. A multivariate calibration strategy based on inverse least squares approach was employed for quantification. [Copyright &y& Elsevier]

Background: Multiple sequence alignments are used to study gene or protein function, phylogenetic relations, genome evolution hypotheses and even gene polymorphisms. Virtually without exception, all available tools focus on conserved segments or residues. Small divergent regions, however, are biologically important for specific quantitative polymerase chain reaction, genotyping, molecular markers and preparation of specific antibodies, and yet have received little attention. As a consequence, they must be selected empirically by the researcher. AlignMiner has been developed to fill this gap in bioinformatic analyses. Results: AlignMiner is a Web-based application for detection of conserved and divergent regions in alignments of conserved sequences, focusing particularly on divergence. It accepts alignments (protein or nucleic acid) obtained using any of a variety of algorithms, which does not appear to have a significant impact on the final results. AlignMiner uses different scoring methods for assessing conserved/divergent regions, Entropy being the method that provides the highest number of regions with the greatest length, and Weighted being the most restrictive. Conserved/divergent regions can be generated either with respect to the consensus sequence or to one master sequence. The resulting data are presented in a graphical interface developed in AJAX, which provides remarkable user interaction capabilities. Users do not need to wait until execution is complete and can.even inspect their results on a different computer. Data can be downloaded onto a user disk, in standard formats. In silico and experimental proof-of-concept cases have shown that AlignMiner can be successfully used to designing specific polymerase chain reaction primers as well as potential epitopes for antibodies. Primer design is assisted by a module that deploys several oligonucleotide parameters for designing primers "on the fly". Conclusions: AlignMiner can be used to reliably detect divergent regions via several scoring methods that provide different levels of selectivity. Its predictions have been verified by experimental means. Hence, it is expected that its usage will save researchers' time and ensure an objective selection of the best-possible divergent region when closely related sequences are analysed. AlignMiner is freely available at http://www.scbi.uma.es/alignminer. [ABSTRACT FROM AUTHOR]

Background: High-throughput automated sequencing has enabled an exponential growth rate of sequencing data. This requires increasing sequence quality and reliability in order to avoid database contamination with artefactual sequences. The arrival of pyro-sequencing enhances this problem and necessitates customisable preprocessing algorithms. Results: SeqTrim has been implemented both as a Web and as a standalone command line application. Already-published and newly-designed algorithms have been included to identify sequence inserts, to remove low quality, vector, adaptor, low complexity and contaminant sequences, and to detect chimeric reads. The availability of several input and output formats allows its inclusion in sequence processing workflows. Due to its specific algorithms, SeqTrim outperforms other pre-processors implemented as Web services or standalone applications. It performs equally well with sequences from EST libraries, SSH libraries, genomic DNA libraries and pyrosequencing reads and does not lead to over-trimming. Conclusions: SeqTrim is an efficient pipeline designed for pre-processing of any type of sequence read, including next-generation sequencing. It is easily configurable and provides a friendly interface that allows users to know what happened with sequences at every pre-processing stage, and to verify pre-processing of an individual sequence if desired. The recommended pipeline reveals more information about each sequence than previously described pre-processors and can discard more sequencing or experimental artefacts. [ABSTRACT FROM AUTHOR]

A central goal of comparative plant ecology is to understand how functional traits vary among species and to what extent this variation has adaptive value. Here we evaluate relationships between four functional traits (seed volume, specific leaf area, wood density, and adult stature) and two demographic attributes (diameter growth and tree mortality) for large trees of 240 tree species from five Neotropical forests. We evaluate how these key functional traits are related to survival and growth and whether similar relationships between traits and demography hold across different tropical forests. There was a tendency for a trade-off between growth and survival across rain forest tree species. Wood density, seed volume, and adult stature were significant predictors of growth and/or mortality. Both growth and mortality rates declined with an increase in wood density. This is consistent with greater construction costs and greater resistance to stem damage for denser wood. Growth and mortality rates also declined as seed volume increased. This is consistent with an adaptive syndrome in which species tolerant of low resource availability (in this case shade-tolerant species) have large seeds to establish successfully and low inherent growth and mortality rates. Growth increased and mortality decreased with an increase in adult stature, because taller species have a greater access to light and longer life spans. Specific leaf area was, surprisingly, only modestly informative for the performance of large trees and had ambiguous relationships with growth and survival. Single traits accounted for 9-55% of the interspecific variation in growth and mortality rates at individual sites. Significant correlations with demographic rates tended to be similar across forests and for phylogenetically independent contrasts as well as for cross-species analyses that treated each species as an independent observation. In combination, the morphological traits explained 41% of the variation in growth rate and 54% of the variation in mortality rate, with wood density being the best predictor of growth and mortality. Relationships between functional traits and demographic rates were statistically similar across a wide range of Neotropical forests. The consistency of these results strongly suggests that tropical rain forest species face similar trade-offs in different sites and converge on similar sets of solutions. [ABSTRACT FROM AUTHOR]

This work describes an easy alkaline lysis method for isolatating bacterial artificial chromosome (BAC) DNA in sufficient quantity and quality for further manipulation without the need to use a kit. The method starts with 10 mL of culture and, by alkaline lysis only, renders up to 150 ng of DNA per milliliter of culture. This BAC DNA was successfully digested with restriction enzymes, sequenced, and subjected to PCR. [ABSTRACT FROM AUTHOR]

There is an urgent need for the development of early identification techniques in olive-trees due to the economic importance of cultivar identification in periods of expansion like now. We have been able to identify 22 olive-tree cultivars using only 10 different, specific, repeatable markers. These markers were designed by the cloning of significant RAPD bands obtained in PCR performed on bulked DNA to retain the genetic variability of each cultivar. Clones were partially or totally sequenced and new primers derived from these sequences were used to obtain Sequence Characterised Amplified Region (SCAR) fragments. We have demonstrated that the use of the 10 SCAR markers is enough to provide a simple, cheap, and reliable procedure to identify 22 geographically related olive-tree cultivars. [ABSTRACT FROM AUTHOR]

Ammonium is assimilated into amino acids through the sequential action of glutamine synthetase (GS) and glutamate synthase (GOGAT) enzymes. This metabolic pathway is driven by energy, reducing power and requires the net supply of 2‐oxoglutarate that can be provided by the reaction catalysed by isocitrate dehydrogenase (IDH). Most studies on the biochemistry and molecular biology of N‐assimilating enzymes have been carried out on annual plant species and the available information on woody models is far more limited. This is in spite of their economic and ecological importance and the fact that nitrogen is a common limiting factor for tree growth. GS, GOGAT and IDH enzymes have been purified from several woody species and their kinetic and molecular properties determined. A number of cDNA clones have also been isolated and characterized. Although the enzymes are remarkably well conserved along the evolutionary scale, major differences have been found in their compartmentation within the cell between angiosperms and conifers, suggesting possible adaptations to specific functional roles. The analysis of the gene expression patterns in a variety of biological situations such as changes in N nutrition, development, biotic or abiotic stresses and senescence, suggest that cytosolic GS plays a central and pivotal role in ammonium assimilation and metabolism in woody plants. The modification of N assimilation efficiency has been recently approached in trees by overexpression of a cytosolic pine GS in poplar. The results obtained, suggest that an increase in cytosolic GS might lead to a global effect on the synthesis of nitrogenous compounds in the leaves, with enhanced vegetative growth of transgenic trees. All these data suggest that manipulation of cytosolic GS may have consequences for plant growth and biomass production. [ABSTRACT FROM AUTHOR]

During evolution, cellular processes leading to the transfer of genetic information failed to send all the mitochondrial genes into the nuclear genome. Two mitochondrial genes are still exclusively located in the mitochondrial genome of all living organisms. They code for two highly hydrophobic proteins: the apocytochrome b and the subunit I of cytochrome oxidase. Assuming that the translocation machinery could not efficiently transport long hydrophobic fragments, we searched for multicopy suppressors of this physical blockage. We demonstrated that overexpression of Pse1p/Kap121p or Kap123p, which belong to the superfamily of karyopherin β proteins, facilitates the translocation of chimeric proteins containing several stretches of apocytochrome b fused to a reporter mitochondrial gene. The effect of PSE1/KAP121 overexpression (in which PSE1 is protein secretion enhancer 1) on mitochondrial import of the chimera is correlated with an enrichment of the corresponding transcript in cytoplasmic ribosomes associated with mitochondria. PSE1/KAP121 overexpression also improves the import of the hydrophobic protein Atm1p, an ABC transporter of the mitochondrial inner membrane. These results suggest that in vivo PSE1/KAP121 overexpression facilitates, either directly or indirectly, the co-translational import of hydrophobic proteins into mitochondria. [ABSTRACT FROM AUTHOR]

Breeding programs are essential for aquaculture. The present study was conducted to investigate the genetic parameters of four growth traits in Senegalese sole. Families were produced by mass spawning (n = 2171 offspring) from an industrial broodstock. Weight, total area, standard length and width (the two latter measured directly on the fish and by image analysis) were determined before the on-growing stage (age ~ 400 d) and at harvest (~800 d) in RAS. Phenotypic data are presented and discussed. Females grew faster than males and a high variation of sex ratios by family was observed. Heritabilities were high for all traits ranging from 0.568 to 0.609 at 400 d and from 0.424 to 0.500 at 800d. The genetic correlations between traits were very high (>0.94) at both ages. Genetic estimates for traits measured in vivo and by digital image analysis were similar. To investigate genomic regions associated with these growth traits, a low-density array using a set of 60 single nucleotide polymorphisms (SNPs) distributed in 17 linkage groups was designed. Using samples from wild fish, a total of 49 assays for SNP analysis were validated. The association analysis was carried out using two fast-growth families and two slow-growth families (n = 279). Two significantly-associated SNPs with most of the traits at both ages were detected. They were located in the general transcription factor 3C polypeptide 4 (gtf3c4) and the mitochondrial fission process protein 1 (mtfp1). Results are of high relevance for genetic breeding programs in this species. • Genetic estimates for four growth traits before on-growing and at harvest in Senegalese sole were determined. • Genetic correlations between growth traits at different ages and between ages were high and positive. • Genetic estimates on image-measured traits are highly consistent with those directly measured on fish. • A low-density array with 49 SNPs widely distributed in the genome was validated. • GWAS analysis identified two markers significantly associated with growth. [ABSTRACT FROM AUTHOR]

Simple Summary: Reduced oxygen availability generates a number of adaptive features across all the animal kingdom, and the goal of this study was targeting the mild-hypoxia driving force for metabolic and muscle transcriptional reprogramming of gilthead sea bream juveniles. Attention was focused on blood metabolic and muscle transcriptomic landmarks before and after exhaustive exercise. Our results after mild-hypoxia conditioning highlighted an increased contribution of lipid metabolism to whole energy supply to preserve the aerobic energy production, a better swimming performance regardless of changes in feed intake, as well as reduced protein turnover and improved anaerobic fitness with the restoration of normoxia. On-growing juveniles of gilthead sea bream were acclimated for 45 days to mild-hypoxia (M-HYP, 40–60% O2 saturation), whereas normoxic fish (85–90% O2 saturation) constituted two different groups, depending on if they were fed to visual satiety (control fish) or pair-fed to M-HYP fish. Following the hypoxia conditioning period, all fish were maintained in normoxia and continued to be fed until visual satiation for 3 weeks. The time course of hypoxia-induced changes was assessed by changes in blood metabolic landmarks and muscle transcriptomics before and after exhaustive exercise in a swim tunnel respirometer. In M-HYP fish, our results highlighted a higher contribution of aerobic metabolism to whole energy supply, shifting towards a higher anaerobic fitness following normoxia restoration. Despite these changes in substrate preference, M-HYP fish shared a persistent improvement in swimming performance with a higher critical speed at exercise exhaustion. The machinery of muscle contraction and protein synthesis and breakdown was also largely altered by mild-hypoxia conditioning, contributing this metabolic re-adjustment to the positive regulation of locomotion and to the catch-up growth response during the normoxia recovery period. Altogether, these results reinforce the presence of large phenotypic plasticity in gilthead sea bream, and highlights mild-hypoxia as a promising prophylactic measure to prepare these fish for predictable stressful events. [ABSTRACT FROM AUTHOR]

Simple Summary: This review is a summary of recent studies of genes, many of them involved in growth and reproduction, which can be used for distinguishing between species of the Anabantoidei suborder of fish, focusing on the Blue Gourami as a model species. This is important in both basic science and aquaculture applications. Markers of genetic variation between species are important for both applied and basic research. Here, various genes of the blue gourami (Trichogaster trichopterus, suborder Anabantoidei, a model labyrinth fish), many of them involved in growth and reproduction, are reviewed as markers of genetic variation. The genes encoding the following hormones are described: kisspeptins 1 and 2, gonadotropin-releasing hormones 1, 2, and 3, growth hormone, somatolactin, prolactin, follicle- stimulating hormone and luteinizing hormone, as well as mitochondrial genes encoding cytochrome b and 12S rRNA. Genetic markers in blue gourami, representing the suborder Anabantoidei, differ from those in other bony fishes. The sequence of the mitochondrial cytochrome c oxidase subunit 1 (COI) gene of blue gourami is often used to study the Anabantoidei suborder. Among the genes involved in controlling growth and reproduction, the most suitable genetic markers for distinguishing between species of the Anabantoidei have functions in the hypothalamic–pituitary–somatotropic axis: pituitary adenylate cyclase-activating polypeptide and growth hormone, and the 12S rRNA gene. [ABSTRACT FROM AUTHOR]

Senegalese sole is an economically important flatfish species in aquaculture and an attractive model to decipher the molecular mechanisms governing the severe transformations occurring during metamorphosis, where retinoic acid seems to play a key role in tissue remodeling. In this study, a robust sole transcriptome was envisaged by reducing the number of assembled libraries (27 out of 111 available), fine-tuning a new automated and reproducible set of workflows for de novo assembling based on several assemblers, and removing low confidence transcripts after mapping onto a sole female genome draft. From a total of 96 resulting assemblies, two "raw" transcriptomes, one containing only Illumina reads and another with Illumina and GS-FLX reads, were selected to provide SOLSEv5.0, the most informative transcriptome with low redundancy and devoid of most single-exon transcripts. It included both Illumina and GS-FLX reads and consisted of 51,348 transcripts of which 22,684 code for 17,429 different proteins described in databases, where 9527 were predicted as complete proteins. SOLSEv5.0 was used as reference for the study of retinoic acid (RA) signalling in sole larvae using drug treatments (DEAB, a RA synthesis blocker, and TTNPB, a RA-receptor agonist) for 24 and 48 h. Differential expression and functional interpretation were facilitated by an updated version of DEGenes Hunter. Acute exposure of both drugs triggered an intense, specific and transient response at 24 h but with hardly observable differences after 48 h at least in the DEAB treatments. Activation of RA signalling by TTNPB specifically increased the expression of genes in pathways related to RA degradation, retinol storage, carotenoid metabolism, homeostatic response and visual cycle, and also modified the expression of transcripts related to morphogenesis and collagen fibril organisation. In contrast, DEAB mainly decreased genes related to retinal production, impairing phototransduction signalling in the retina. A total of 755 transcripts mainly related to lipid metabolism, lipid transport and lipid homeostasis were altered in response to both treatments, indicating non-specific drug responses associated with intestinal absorption. These results indicate that a new assembling and transcript sieving were both necessary to provide a reliable transcriptome to identify the many aspects of RA action during sole development that are of relevance for sole aquaculture. [ABSTRACT FROM AUTHOR]

The authors discuss second-growth forests in the Amazon River Basin on topics including the decline of the net carbon uptake of Amazonian old-growth forests, the role of regenerating forests as carbon sinks, and net carbon sequestration rates after timber extraction.

Familial predisposition is among the major genetic risk factors for non-contact musculoskeletal tissue injuries. Personal genome sequence shows that different polymorphism profiles may account for the number and the degree of injuries and the recovery time. Genotyping studies allow investigation into genome factors with potential impact on pathogenesis of non-contact ligament injuries. We have studied a family with twin sibling males surgically diagnosed of an anterior cruciate ligament non-contact rupture and non-affected progenitors (father and mother) were subjected to whole exome sequencing (WES) analysis. WES analysis previously carried out on 16 individuals, without ACL injury medical records, were also included in this study for single nucleotide variants (SNVs) and small insertions and deletions detection (indels), variant filtering and to prioritize variants relative to the disease. WES analysis to identify SNVs and indels was performed using open web-based bioinformatics tools. A set of 11 new variants shared by family members can be associated to ACL non-contact injury, including SerpinA11, ARSI, NOCHT4, EPB41, FDFT1, POMC, KIF26A, OLFML2B, ATG7, FAH and WDR6. All of them, except ATG7 and WDR6, have shown a damaging predictive pattern by combinatorial standard predictive scores. In combination to the identified SNVs of EPB41 and SerpinA11 genes, ACTL7A gene showed a predicted deleterious variant reinforcing the idea these variants impact on of fibroblast-like cells deformability and ECM misbalance, Differential gene expression and RNA sequencing analysis will help to understand the combined participation of these protein coding genes in ACL non-contact injuries. [ABSTRACT FROM AUTHOR]

Introduction Exposure to both prenatal stress (PS) and chronic stress in later life is associated with the development of mood disorders like depression. PS has been found to be related with alterations in the fetal hypothalamo-pituitary-adrenal (HPA) axis, which, in turn, may influence the serotonergic (5-HT) system. 5-HT may also be involved in depression promoted by chronic stress in later life. Therefore, disturbances in the 5-HT system may play an important role in the pathology of depression. In the present study we examined the effects of prenatal restraint stress and/or chronic mild stress (CMS) on 5-HT and tryptophan hydroxylase 2 (TPH2) immunoreactivity within the dorsal raphe nucleus, the hippocampus and the prefrontal cortex of adult male and female Sprague-Dawley rats. Methodology Pregnant rats were individually restrained in transparent plastic cylinders whilst being exposed to bright light three times a day during the last week of pregnancy. For CMS, two random stressors, e.g. empty cage or flashing lights, each lasting for three hours, were applied for a period of three weeks. The effects of PS and/or CMS on 5-HT and TPH2 were analyzed by means of immunohistochemistry and the NIH ImageJ software. Obtained values were statistically analyzed using Two-way ANOVA and post-hoc LSD. Results The data show that PS and chronic stress in later life affect the rat 5-HT system in a region- and sex-specific way. In males, a significant interaction of PS and CMS on the density of positive 5-HT neurons in the dorsal raphe nucleus was observed. Further, a tendency towards increased hippocampal 5-HT immunoreactivity, and significantly elevated 5-HT intensity in the prefrontal cortex were found. In females, stress did not result in any alterations. Concerning TPH2, a PS and CMS interaction was observed in the hippocampus within both sexes. Conclusion PS and CMS alter the 5-HT system in males, with increased hippocampal and prefrontal cortex 5-HT immunoreactivity and reduced levels in the raphe, which might represent compensatory mechanisms. In females no alteration was found indicating a higher susceptibility in males. Regarding TPH2, both sexes were affected to a lesser extent. CMS partially normalized some effects of PS. The observed changes may represent a morphological basis of PS- and/or CMS-related emotional disturbances. [ABSTRACT FROM AUTHOR]