Estudi tècnic-econòmic de l'adaptació a la Llei de Benestar Animal d'una explotació porcina de la comarca d'Osona i posterior valoració de la seva implementació

Des de la publicació de la Llei de Benestar Animal RD 1135/ 2002, de 31 d’octubre que regula els tres aspectes clau de la producció porcina‐ sistemes d’estabulació i construccions permeses; formació obligatòria dels ramaders i maneig dels animals‐ totes les granges de nova construcció han estat projectades seguint els seus preceptes. Mentre que les granges existents han hagut d’adaptar‐hi les seves instal∙lacions sota amenaça de tancament si no es complien els requisits. L’objectiu d’aquest treball ha estat l’elaboració de l’estudi tècnic‐ econòmic per a l’adaptació a la Llei de Benestar Animal d’una explotació porcina de la comarca d’Osona. Després d’avaluar la situació inicial de les instal∙lacions, establir les línies de previsió de creixement i analitzar els avantatges i inconvenients de cada sistema i model, s’opta pel sistema d’alimentació electrònic model Nedap en grup dinàmic de truges sobre sòl amb jaç de palla. Una decisió no massa extesa en granges del sud d’Europa on se sol preferir l’slat de formigó i/o superfície pavimentada, però imprescindible perquè aquesta explotació pogués aconseguir una millor gestió de les dejeccions ramaderes i assegurar una millora agronòmica de les terres de conreu. El treball conclou amb el seguiment del desenvolupament de l’alternativa escollida i una valoració dels canvis derivats de l’adaptació al Benestar Animal després del primer any i mig en ús. Les millores obtingudes en termes de maneig, funcionament de la granja, salut dels animals i índex productius són remarcables. D’una banda, cal destacar el fet de tenir un nombre de coixeres molt menor respecte les granges amb superfície dura, així com la facilitat i rapidesa en els parts degut al benestar. De l’altra,la reducció del volum de purí, l’increment de la fracció sòlida i la disponibilitat de compost per al camp. Per això, no és exagerat observar el jaç amb palla com una de les solucions més adequades i equilibrades per aquesta explotació que combina boví, porcí i terra per a la producció de cultius.

Functional and evolutionary analysis of DXL1, a non-essential gene encoding a 1-deoxy-D-xylulose 5-phosphate synthase like protein in Arabidopsis thaliana

The synthesis of 1-deoxy-D-xylulose 5-phosphate (DXP), catalyzed by the enzyme DXP synthase (DXS), represents a key regulatory step of the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway for isoprenoid biosynthesis. In plants DXS is encoded by small multigene families that can be classified into, at least, three specialized subfamilies. Arabidopsis thaliana contains three genes encoding proteins with similarity to DXS, including the well-known DXS1/CLA1 gene, which clusters within subfamily I. The remaining proteins, initially named DXS2 and DXS3, have not yet been characterized. Here we report the expression and functional analysis of A. thaliana DXS2. Unexpectedly, the expression of DXS2 failed to rescue Escherichia coli and A. thaliana mutants defective in DXS activity. Coherently, we found that DXS activity was negligible in vitro, being renamed as DXL1 following recent nomenclature recommendation. DXL1 is targeted to plastids as DXS1, but shows a distinct expression pattern. The phenotypic analysis of a DXL1 defective mutant revealed that the function of the encoded protein is not essential for growth and development. Evolutionary analyses indicated that DXL1 emerged from DXS1 through a recent duplication apparently specific of the Brassicaceae lineage. Divergent selective constraints would have affected a significant fraction of sites after diversification of the paralogues. Furthermore, amino acids subjected to divergent selection and likely critical for functional divergence through the acquisition of a novel, although not yet known, biochemical function, were identified. Our results provide with the first evidences of functional specialization at both the regulatory and biochemical level within the plant DXS family.

CDK4 is an essential insulin effector in adipocytes.

Insulin resistance is a fundamental pathogenic factor that characterizes various metabolic disorders, including obesity and type 2 diabetes. Adipose tissue contributes to the development of obesity-related insulin resistance through increased release of fatty acids, altered adipokine secretion, and/or macrophage infiltration and cytokine release. Here, we aimed to analyze the participation of the cyclin-dependent kinase 4 (CDK4) in adipose tissue biology. We determined that white adipose tissue (WAT) from CDK4-deficient mice exhibits impaired lipogenesis and increased lipolysis. Conversely, lipolysis was decreased and lipogenesis was increased in mice expressing a mutant hyperactive form of CDK4 (CDK4R24C). A global kinome analysis of CDK4-deficient mice following insulin stimulation revealed that insulin signaling is impaired in these animals. We determined that insulin activates the CCND3-CDK4 complex, which in turn phosphorylates insulin receptor substrate 2 (IRS2) at serine 388, thereby creating a positive feedback loop that maintains adipocyte insulin signaling. Furthermore, we found that CCND3 expression and IRS2 serine 388 phosphorylation are increased in human obese subjects. Together, our results demonstrate that CDK4 is a major regulator of insulin signaling in WAT.

Apical access and closure devices for transapical transcatheter heart valve procedures.

The majority of transcatheter aortic valve implantations, structural heart procedures and the newly developed transcatheter mitral valve repair and replacement are traditionally performed either through a transfemoral or a transapical access site, depending on the presence of severe peripheral vascular disease or anatomic limitations. The transapical approach, which carries specific advantages related to its antegrade nature and the short distance between the introduction site and the cardiac target, is traditionally performed through a left anterolateral mini-thoracotomy and requires rib retractors, soft tissue retractors and reinforced apical sutures to secure, at first, the left ventricular apex for the introduction of the stent-valve delivery systems and then to seal the access site at the end of the procedure. However, despite the advent of low-profile apical sheaths and newly designed delivery systems, the apical approach represents a challenge for the surgeon, as it has the risk of apical tear, life-threatening apical bleeding, myocardial damage, coronary damage and infections. Last but not least, the use of large-calibre stent-valve delivery systems and devices through standard mini-thoracotomies compromises any attempt to perform transapical transcatheter structural heart procedures entirely percutaneously, as happens with the transfemoral access site, or via a thoracoscopic or a miniaturised video-assisted percutaneous technique. During the past few years, prototypes of apical access and closure devices for transapical heart valve procedures have been developed and tested to make this standardised successful procedure easier. Some of them represent an important step towards the development of truly percutaneous transcatheter transapical heart valve procedures in the clinical setting.

Functional EF-hands in neuronal calcium sensor GCAP2 determine its phosphorylation state and subcellular distribution in vivo, and are essential for photoreceptor cell integrity

The neuronal calcium sensor proteins GCAPs (guanylate cyclase activating proteins) switch between Ca2+-free and Ca2+-bound conformational states and confer calcium sensitivity to guanylate cyclase at retinal photoreceptor cells. They play a fundamental role in light adaptation by coupling the rate of cGMP synthesis to the intracellular concentration of calcium. Mutations in GCAPs lead to blindness. The importance of functional EF-hands in GCAP1 for photoreceptor cell integrity has been well established. Mutations in GCAP1 that diminish its Ca2+ binding affinity lead to cell damage by causing unabated cGMP synthesis and accumulation of toxic levels of free cGMP and Ca2+. We here investigate the relevance of GCAP2 functional EF-hands for photoreceptor cell integrity. By characterizing transgenic mice expressing a mutant form of GCAP2 with all EF-hands inactivated (EF(-)GCAP2), we show that GCAP2 locked in its Ca2+-free conformation leads to a rapid retinal degeneration that is not due to unabated cGMP synthesis. We unveil that when locked in its Ca2+-free conformation in vivo, GCAP2 is phosphorylated at Ser201 and results in phospho-dependent binding to the chaperone 14-3-3 and retention at the inner segment and proximal cell compartments. Accumulation of phosphorylated EF(-)GCAP2 at the inner segment results in severe toxicity. We show that in wildtype mice under physiological conditions, 50% of GCAP2 is phosphorylated correlating with the 50% of the protein being retained at the inner segment. Raising mice under constant light exposure, however, drastically increases the retention of GCAP2 in its Ca2+-free form at the inner segment. This study identifies a new mechanism governing GCAP2 subcellular distribution in vivo, closely related to disease. It also identifies a pathway by which a sustained reduction in intracellular free Ca2+ could result in photoreceptor damage, relevant for light damage and for those genetic disorders resulting in 'equivalent-light'' scenarios.

Type II aromatic polyketide biosynthetic tailoring enzymes: diversity and adaptation in Streptomyces secondary metabolism.

Members of the bacterial genus Streptomyces are well known for their ability to produce an exceptionally wide selection of diverse secondary metabolites. These include natural bioactive chemical compounds which have potential applications in medicine, agriculture and other fields of commerce. The outstanding biosynthetic capacity derives from the characteristic genetic flexibility of Streptomyces secondary metabolism pathways: i) Clustering of the biosynthetic genes in chromosome regions redundant for vital primary functions, and ii) the presence of numerous genetic elements within these regions which facilitate DNA rearrangement and transfer between non-progeny species. Decades of intensive genetic research on the organization and function of the biosynthetic routes has led to a variety of molecular biology applications, which can be used to expand the diversity of compounds synthesized. These include techniques which, for example, allow modification and artificial construction of novel pathways, and enable gene-level detection of silent secondary metabolite clusters. Over the years the research has expanded to cover molecular-level analysis of the enzymes responsible for the individual catalytic reactions. In vitro studies of the enzymes provide a detailed insight into their catalytic functions, mechanisms, substrate specificities, interactions and stereochemical determinants. These are factors that are essential for the thorough understanding and rational design of novel biosynthetic routes. The current study is a part of a more extensive research project (Antibiotic Biosynthetic Enzymes; www.sci.utu.fi/projects/biokemia/abe), which focuses on the post-PKS tailoring enzymes involved in various type II aromatic polyketide biosynthetic pathways in Streptomyces bacteria. The initiative here was to investigate specific catalytic steps in anthracycline and angucycline biosynthesis through in vitro biochemical enzyme characterization and structural enzymology. The objectives were to elucidate detailed mechanisms and enzyme-level interactions which cannot be resolved by in vivo genetic studies alone. The first part of the experimental work concerns the homologous polyketide cyclases SnoaL and AknH. These catalyze the closure of the last carbon ring of the tetracyclic carbon frame common to all anthracycline-type compounds. The second part of the study primarily deals with tailoring enzymes PgaE (and its homolog CabE) and PgaM, which are responsible for a cascade of sequential modification reactions in angucycline biosynthesis. The results complemented earlier in vivo findings and confirmed the enzyme functions in vitro. Importantly, we were able to identify the amino acid -level determinants that influence AknH and SnoaL stereoselectivity and to determine the complex biosynthetic steps of the angucycline oxygenation cascade of PgaE and PgaM. In addition, the findings revealed interesting cases of enzyme-level adaptation, as some of the catalytic mechanisms did not coincide with those described for characterised homologs or enzymes of known function. Specifically, SnoaL and AknH were shown to employ a novel acid-base mechanism for aldol condenzation, whereas the hydroxylation reaction catalysed by PgaM involved unexpected oxygen chemistry. Owing to a gene-level fusion of two ancestral reading frames, PgaM was also shown to adopt an unusual quaternary sturucture, a non-covalent fusion complex of two alternative forms of the protein. Furthermore, the work highlighted some common themes encountered in polyketide biosynthetic pathways such as enzyme substrate specificity and intermediate reactivity. These are discussed in the final chapters of the work.

Active sulphide mine tailings impoundments as sources of contaminated drainage: controlling factors, methods of characterisation and geochemical constraints for mitigation

Low quality mine drainage from tailings facilities persists as one of the most significant global environmental concerns related to sulphide mining. Due to the large variation in geological and environmental conditions at mine sites, universal approaches to the management of mine drainage are not always applicable. Instead, site-specific knowledge of the geochemical behaviour of waste materials is required for the design and closure of the facilities. In this thesis, tailings-derived water contamination and factors causing the pollution were investigated in two coeval active sulphide mine sites in Finland: the Hitura Ni mine and the Luikonlahti Cu-Zn-Co-Ni mine and talc processing plant. A hydrogeochemical study was performed to characterise the tailingsderived water pollution at Hitura. Geochemical changes in the Hitura tailings were evaluated with a detailed mineralogical and geochemical investigation (solid-phase speciation, acid mine drainage potential, pore water chemistry) and using a spatial assessment to identify the mechanisms of water contamination. A similar spatial investigation, applying selective extractions, was carried out in the Luikonlahti tailings area for comparative purposes (Hitura low-sulphide tailings vs. Luikonlahti sulphide-rich tailings). At both sites, hydrogeochemistry of tailings seepage waters was further characterised to examine the net results of the processes observed within the impoundments and to identify constraints for water treatment. At Luikonlahti, annual and seasonal variation in effluent quality was evaluated based on a four-year monitoring period. Observations pertinent to future assessment and mine drainage prevention from existing and future tailings facilities were presented based on the results. A combination of hydrogeochemical approaches provided a means to delineate the tailings-derived neutral mine drainage at Hitura. Tailings effluents with elevated Ni, SO₄ ^2- and Fe content had dispersed to the surrounding aquifer through a levelled-out esker and underneath the seepage collection ditches. In future mines, this could be avoided with additional basal liners in tailings impoundments where the permeability of the underlying Quaternary deposits is inadequate, and with sufficiently deep ditches. Based on the studies, extensive sulphide oxidation with subsequent metal release may already initiate during active tailings disposal. The intensity and onset of oxidation depended on e.g. the Fe sulphide content of the tailings, water saturation level, and time of exposure of fresh sulphide grains. Continuous disposal decreased sulphide weathering in the surface of low-sulphide tailings, but oxidation initiated if they were left uncovered after disposal ceased. In the sulphide-rich tailings, delayed burial of the unsaturated tailings had resulted in thick oxidized layers, despite the continuous operation. Sulphide weathering and contaminant release occurred also in the border zones. Based on the results, the prevention of sulphide oxidation should already be considered in the planning of tailings disposal, taking into account the border zones. Moreover, even lowsulphide tailings should be covered without delay after active disposal ceases. The quality of tailings effluents showed wide variation within a single impoundment and between the two different types of tailings facilities assessed. The affecting factors included source materials, the intensity of weathering of tailings and embankment materials along the seepage flow path, inputs from the process waters, the water retention time in tailings, and climatic seasonality. In addition, modifications to the tailings impoundment may markedly change the effluent quality. The wide variation in the tailings effluent quality poses challenges for treatment design. The final decision on water management requires quantification of the spatial and seasonal fluctuation at the site, taking into account changes resulting from the eventual closure of the impoundment. Overall, comprehensive hydrogeochemical mapping was deemed essential in the identification of critical contaminants and their sources at mine sites. Mineralogical analysis, selective extractions, and pore water analysis were a good combination of methods for studying the weathering of tailings and in evaluating metal mobility from the facilities. Selective extractions with visual observations and pH measurements of tailings solids were, nevertheless, adequate in describing the spatial distribution of sulphide oxidation in tailings impoundments. Seepage water chemistry provided additional data on geochemical processes in tailings and was necessary for defining constraints for water treatment.

Composition and antifungal activity of essential oils from Piper aduncum, Piper arboreum and Piper tuberculatum

The composition of essential oils from leaves, stems and fruits of Piper aduncum, P. arboreum and P. tuberculatum was examined by means of GC-MS and antifungal assay. There was a predominance of monoterpenes in P. aduncum and P. tuberculatum and of sesquiterpenes in P. arboreum. P. aduncum showed the richest essential oil composition, including linalool. The essential oils from fruits of P. aduncum and P. tuberculatum showed the highest antifungal activity with the MIC of 10 µg as determined against Cladosporium cladosporioides and C. sphaerospermum, respectively. This is the first report of the composition of essential oils from P. tuberculatum.

Essential factors for control of the equilibrium in the reversible rearrangement of M3N@Ih-C80 fulleropyrrolidines: Exohedral functional groups versus endohedral metal clusters

The effects of exohedral moieties and endohedral metal clusters on the isomerization of M3N@Ih-C80 products from the Prato reaction through [1,5]-sigmatropic rearrangement were systematically investigated by using three types of fulleropyrrolidine derivatives and four different endohedral metal clusters. As a result, all types of derivatives provided the same ratios of the isomers for a given trimetallic nitride template (TNT) as the thermodynamic products, thus indicating that the size of the endohedral metal clusters inside C80 was the single essential factor in determining the equilibrium between the [6,6]-isomer (kinetic product) and the [5,6]-isomer. In all the derivatives, the [6,6]- and [5,6]-Prato adducts with larger metal clusters, such as Y3N and Gd3N, were equally stable, which is in good agreement with DFT calculations. The reaction rate of the rearrangement was dependent on both the substituent of exohedral functional groups and the endohedral metal-cluster size. Further DFT calculations and 13C NMR spectroscopic studies were employed to rationalize the equilibrium in the rearrangement between the [6,6]- and [5,6]-fulleropyrrolidines

Chemical composition and antimicrobial activity of the essential oil from Aeolanthus suaveolens Mart. ex Spreng

The essential oils from leaves (sample A) and flowers (sample B) of Aeolanthus suaveolens Mart. ex Spreng were obtained by hydrodistillation and analyzed by GC, GC-MS, and chiral phase gas chromatography (CPGC). Six compounds have been identified from the essential oils, representing ca 94.3 and 93% of the oils corresponding to samples A and B, respectively. The major constituents of samples A and B essential oils were respectively, linalool (34.2%/34.9%), (-)-massoialactone (25.9%/17.0%) and (E)-beta-farnesene (25.4%/29.1%). The enantiomeric distribution of the monoterpene linalool was established by analysis on heptakis- (6-O-methyl-2,3-di-O-pentyl)-beta-cyclodextrin capillary column. The antimicrobial activity of the essential oil from leaves and isolated compounds was also evaluated.

Realització d'una aplicació de gestió de xarxes per a la xarxa sensefils de Manresa GuifiBages

En els últims anys, la popularitat de les xarxes sensefils (WIFI) ha anat en augment a un ritme incansable. Des de petits aparells instal•lats a les cases amb aquesta tecnologia com a complement dels routers d’accés a internet instal•lats per diverses companyies, fins a empreses fent petits desplegaments per comunicar entre si les seves seus. Al marge d’aquests escenaris, s’ha produït un fenomen social d’acolliment d’aquesta tecnologia a nivell mundial, en forma del que coneixem com a xarxes ciutadanes / xarxes lliures / xarxes socials. Aquestes xarxes han estat possibles gràcies a diverses raons que han fet assequible a col•lectius de persones, tant els aparells com els coneixements necessaris per dur a terme aquestes actuacions. Dintre d’aquest marc, al Bages, concretament a Manresa, es va començar a desenvolupar una d’aquestes xarxes. Les decisions d’aquesta xarxa d’utilitzar exclusivament hardware i software de codi obert, i determinats aspectes tècnics de la xarxa, ha comportat que la xarxa fos incompatible amb algunes de les aplicacions de gestió de xarxes existents desenvolupades per comunicats com gufi.net a Osona. És per això que per garantir el creixement, la supervivència i l’èxit d’aquesta xarxa en el temps, és indispensable poder comptar amb una eina de gestió que s’adigui a les característiques de GuifiBages. L’objectiu principal d’aquest treball és dotar a la xarxa GuifiBages de les eines necessàries per poder gestionar tota la informació referent a l’estructura de la seva xarxa, tant per facilitar l’accés a nous usuaris sense molts coneixements tècnics, com per facilitar nous desplegaments / reparacions / modificacions de la xarxa d’una manera automàtica. Com a conclusió d’aquest treball, podem afirmar que les avantatges que proporciones tecnologies com Plone, faciliten enormement la creació d’aplicacions de gestió de continguts en entorn web. Alhora, l’ús de noves tècniques de programació com AJAX o recursos com els que ofereix Google, permeten desenvolupar aplicacions web que no tenen res a envejar al software tradicional. D’altra banda, voldríem destacar l’ús exclusiu de programari lliure tant en els paquets de software necessaris pel desenvolupament, com en el sistema operatiu i programes dels ordinadors on s’ha dut a terme, demostrant que es poden desenvolupar sistemes de qualitat sense dependre de programari privatiu.

Essential oils from leaves of cryptocarya spp from the atlantic rain forest

The essential oils from leaves of four Cryptocarya spp endemic in the Brazilian Atlantic rain forest were obtained by hydrodistillation and shown by GC-MS analysis to contain mono and sesquiterpenes. The major components of the oil of Cryptocarya moschata were linalool (34.3%), a-terpinene (17.0%), g-terpinene (10.4%), 1,8-cineole (5.8%) and trans-ocimene (4.8%), whilst those of C. botelhensis were a-pinene (22.7%), b-pinene (9.2%), trans-verbenol (8.4%), trans-pinocarveol (5.5%) and myrtenal (5.4%). The principal compounds of C. mandioccana oil were b-caryophyllene (13.8%), spathulenol (10.2%), caryophyllene oxide (7.8%), d-cadinene (6.9%) and bicyclogermacrene (6.4%), whilst those of C. saligna were germacrene D (15.5%), bicyclogermacrene (13.8%), spathulenol (11.8%) and germacrene B (5.7%).

Chemical composition and antimicrobial activity of the essential oil of Hyptis pectinata (l.) Poit.

Essential oil was extracted from leaves of Hyptis pectinata using hydrodistillation, and its composition determined using GC-FID and GC-MS. Chemical analysis showed that there was a predominance of sesquiterpenes, of which β-caryophyllene (18.34%), caryophyllene oxide (18.00%) and calamusenone (24.68%) were measured for the first time in the genus Hyptis. Twenty-one compounds were identified, and calamusenone was isolated using preparative thin layer chromatography with a silica gel plate (60 PF254). The minimal inhibitory concentration (MIC) and minimal microbicidal concentration (MMC) were determined for various pathogenic microorganisms. H. pectinata oil was most effective against Gram (+) bacteria and yeasts.

Antimicrobial and antileishmanial activity of essential oil from the leaves of Annona foetida (Annonaceae)

bicyclogermacrene (35.12%), (E)-caryophyllene (14.19%) and α-copaene (8.19%). The antimicrobial and antileishmanial activities were investigated. The oil showed potent antimicrobial activity against Candida albicans and Rhodococcus equi. The oil also showed significant antileishmanial activity, giving the best results against Leishmania guyanensis. A preliminary cytotoxicity assay for this oil was carried out on hamster and mice (Balb/c) peritoneal macrophages. The results obtained were similar to pentamidine and considered not to be cytotoxic to macrophages.