A new subfamily of fungal subtilases: structural and functional analysis of a Pleurotus ostreatus member

Pleurotus ostreatus produces several extracellular proteases which are believed to be involved in the regulation of the ligninolytic activities of this fungus. Recently, purification and characterization of the most abundant P. ostreatus extracellular protease (PoSl) have been reported. The sequence of the posl gene and of the corresponding cDNA has been determined, allowing the identification of its pre- and pro-sequences. A mature protein sequence has been verified by mass spectrometry mapping, the N-glycosylation sites have been identified and the glycosidic moieties characterized. Mature PoSl shows a cleaved peptide bond in the C-terminal region, which remains associated with the catalytic domain in a non-covalent complex. Reported results indicate that this enzyme is involved in the activation of other P. ostreatus secreted proteases, thus suggesting its leading role in cascade activation mechanisms. Analyses of the PoSl sequence by homology search resulted in the identification of a DNA sequence encoding a new protease, homologous to PoSl, in the Phanerochaete chrysosporium genome. A new subgroup of subtilisin-like proteases, belonging to the pyrolysin family, has been defined, which includes proteases from ascomycete and basidiomycete fungi

Assessing the biomass of shrubs typical of Mediterranean pre-forest communities

One of the most outstanding issues of current environmental research is the need of reliable assessments of carbon stock (i.e. above-ground (a.g.) and below-ground biomass, deadwood, litter, and organic soil matter) within forest ecosystems. Although shrub vegetation plays an important role in accumulating carbon in many Mediterranean environments, there is still very little knowledge on the carbon they store. In this article, we analyze the a.g. carbon stock of several Mediterranean shrubby communities in Sicily (Italy), dominated by Pistacia lentiscus, Chamaerops humilis, Euphorbia dendroides and Spartium junceum. Plant samples for each species were selected, and morphometric attributes (height, crown diameter, diameter at breast height, etc.) were measured to establish models for estimating a.g. biomass. Dendrometric data were also used to support the assessment of shrub biomass at community level by coupling them with the coverage of the considered woody species estimated from several hundreds of phytosociological releve´s carried out on pre-forest Sicilian communities. Although the results of such an approach have to be considered as rough assessments, the trial proves interesting, given the lack of information on stand biomass of pre-forest vegetation within the Mediterraean region.L'articolo è disponibile sul sito dell'editore http://www.tandfonline.co

Valore storico, culturale e paesaggistico delle “muracche” nel Bosco Ficuzza

Il vasto complesso forestale indicato come Bosco della Ficuzza ebbe origine quando Ferdinando IV di Borbone, costretto dagli avvenimenti tumultuosi del 1798 a fuggire da Napoli si trasferì in Sicilia. Essendo un grande appassionato di caccia, il sovrano fece costituire diverse tenute di caccia; così sorsero le tenute reali della Favorita a Palermo e quella di Ficuzza. Il Re migliorò la viabilità per giungere a Ficuzza, e poi quella interna al bosco. Nel 1802 diede l’incarico prima all’architetto palermitano Venazio Marvuglia e poi all’architetto Carlo Chenchi di edificare la Real Casina di Caccia, dove dimorò per lunghi anni, praticando, tra tutti i suoi preferiti svaghi, la caccia e la pesca. A Ficuzza, Ferdinando IV si adoperò a costituire una delle sue più grandi riserve di caccia. Così vennero riuniti i feudi di Lupo, Ficuzza e Cappelliere, costituiti i confini con “muracche” (muraglie in pietra) e pilieri recanti la scritta “R.R.“, Reali Riserve, impose norme restrittive alla libera caccia. Altresì, il re dotò il bosco di una ben distribuita rete viaria e si adoperò alla cura dei boschi, al ripopolamento della selvaggina, facendo arrivare da fuori cinghiali, daini e cervi. Inoltre, fece costruire in vari posti del bosco rifugi per i guardiacaccia, masserie e ricoveri per gli armenti, avendo costituito grossi allevamenti di bovini e ovini, abbeveratoi e altre “infrastrutture” al servizio della caccia come il “pulpito del re”, ricavato in un grosso masso di arenaria dove il re si appostava per sparare a lepri, cinghiali e daini che i battitori stanavano. Altra struttura degna di menzione è la “pescheria del Gorgo del Drago”, uno stagno alimentato da una sorgente perenne dove il re, durante il riposo venatorio per il ripopolamento della selvaggina, si dilettava a pescare, ospite nella adiacente casa, oggi ridotta ad un rudere. Le muracche erano state costruite con massi di varia dimensione accatastati in modo da formare dei parallelepipedi di larghezza di circa 1 m, e alte fino a 2 m, a formare estese muraglie lungo i confini delle tenute di caccia reali. Una sorta di recinzioni che servivano a proteggere le aree destinate al ripopolamento della selvaggina ma anche i boschi, tanto cari al re, soggetti agli interventi selvicolturali. Infatti, il re al suo primo sopralluogo a Ficuzza, scrisse nel suo diario di bordo “... i boschi sono ampi e ricchi di selvaggina, ma mal tenuti e troppo sfruttati”. Ecco perché, appena stabilitosi a Ficuzza, il Re iniziò ad adoperarsi per ricostituire i boschi degradati e troppo sfruttati, proteggendoli dal pascolo e da ogni pericolo con le muracche. Lungo le muracche erano presenti dei passaggi delimitati da cancelli, usati per il transito dei boscaioli e della selvaggina. Nei vertici delle muracche erano presenti i pilieri in pietra che, oltre alla scritta “R.R.”, riportavano sopra quest’ultima il simbolo della corona reale. L’opera meritoria di Ferdinando IV di Borbone è stata decisiva nella destinazione della “Foresta nazionale inalienabile di Ficuzza” come “Stazione climatica” ai sensi della Legge n. 535 del 1901 del Regno d’Italia. Delle muracche originarie oggi sono rimaste solo le vestigia coperte da muschi e intrigate nella vegetazione. L’Ente gestore della Riserva ha ripristinato solo una piccola parte degli antichi manufatti a scopo dimostrativo della originaria forma strutturale. L’obiettivo di questo lavoro è stato quello di realizzare un inventario dell’attuale consistenza e stato di manutenzione delle muracche e di tutte le altre infrastrutture ancora esistenti al fine di ricostruire il loro originale sviluppo all’interno della foresta. Tale inventario, oltre ad essere importante dal punto di vista storico, culturale e paesaggistico, può contribuire alla ricostruzione storica della gestione forestale dei boschi di Ficuzza e, al tempo stesso, può avere un ruolo chiave nella definizione del particellare ai fini della redazione del piano di gestione forestale della riserva. Può inoltre essere la base per avviare un serio progetto di recupero di questo patrimonio indissolubilmente legato alla storia umana e naturale di Ficuzza.The vast wooded area known as “Bosco Ficuzza” originated when Ferdinando IV Borbone, forced by the tumultuous events in 1798 to escape from Naples, moved to Sicily. The King, being a great hunting enthusiast, established several hunting estates; thus the royal estates of the Favorita in Palermo and that of Ficuzza rose. The King improved the viability to reach Ficuzza, and then those inside the forest. In 1802 he was commissioned to the venetian architect Venazio Marvuglia and then to the architect Carlo Chenchi to build the “Real Casina di Caccia”, where he lived for many years, practicing, among all his favorite leisure activities, hunting and fishing. In Ficuzza, Ferdinando IV worked to constitute one of his largest hunting reserves. Thus the fiefs of Lupo, Ficuzza and Cappelliere were reunited, the boundaries constituted with “muracche”" (stone walls) and “pilieri” (stone stacks) bearing the inscription "R.R.", Reali Riserve (i.e. Royal Reserves), and he imposed restrictive rules to free hunting. Also, the King endowed the forest with a well-distributed road network and he worked to the care of the woods, to repopulate the game species, by bringing out wild boar, fallow deer and deer. Moreover, in various places of the forest, he had it built shelters for the keepers, farms and shelters for livestock, having formed large cattle and sheep farms, drinking troughs and other infrastructures at the service of hunting such as the "The King's pulpit", obtained from a large sandstone boulder where the King used to shoot hares, wild boars and fallow deer that the beaters used to drive out. Another structure worthy of mention is the “fishmonger of the Gorgo del Drago”, a pond fed by a perennial spring where the King, during the hunting rest for the repopulation of the game species, was delighted to fish, guest in the adjacent house, today reduced to a ruin. The muracche had been built with boulders of various dimensions stacked so as to form parallelepipeds with a width of about 1 m, and up to 2 m tall, to form extensive walls along the boundaries of royal hunting estates. They were a kind of fences that served to protect the areas destined to the repopulation of the game species but also the woods, so dear to the King, subjected to silvicultural interventions. In fact, the King at his first visit to Ficuzza, wrote in his logbook “... the woods are wide and rich in game species, but poorly managed and too much exploited”. This is why, as soon as he settled in Ficuzza, the King began to work to recover the degraded and over-exploited forests, protecting them from grazing and from any other danger with the muracche. Along the muracche there were passages delimited by gates, used for the transit of woodcutters and game species. At the top of the muracche there were stone stacks which, in addition to the inscription “R.R.”, had the symbol of the royal crown above it. The meritorious work of Ferdinand IV of Bourbon was decisive in the destination of the "Inalienable National Forest of Ficuzza" as "Climate Station" in accordance with the Law n. 535 of 1901 of the Kingdom of Italy. Of the original muracche today there are only the remains covered by mosses and intrigued in the vegetation. The managing body of the Reserve has restored only a small part of the ancient artifacts for demonstration purposes of the original structural form. The objective of this work is to create an inventory of the current consistency and state of maintenance of the muracche and of all the other infrastructures still existing in order to reconstruct their original development within the forest. This inventory, in addition to being important from an historical, cultural and landscape point of view, can contribute to the historical reconstruction of the forest management of the Ficuzza woods and, at the same time, can play a key role in defining particles for the purposes of drawing up the forest management plan of the reserve. It can also be the basis for starting a serious project to recover this heritage inextricably linked to the human and natural history of Ficuzza

Targeting IL-6 signalling in early rheumatoid arthritis is followed by Th1 and Th17 suppression and Th2 expansion

OBJECTIVES: To investigate the in vitro and ex-vivo effect of IL-6 inhibition on the balance between Th1, Th2, Th17 and Treg cells. METHODS: Ten consecutive adult patients with active early rheumatoid arthritis (ERA) and ten healthy volunteers were included in the study. The percentages of Th1, Th2, Th17 and Treg cells were analysed by flow cytometry in the peripheral blood mononuclear cells obtained from controls and from RA patients at the time of first evaluation and just before the third TCZ infusion. The in vitro effect of TCZ on the different subsets of CD4+ T cells and the expression levels of Th1, Th2, Th17 and Treg-related cytokines was also assessed. RESULTS: Treatment with TCZ, both ex vivo and in vitro, resulted in a significant reduction of the percentage of Th1, Th17 and Treg cells with a concomitant significant increase of Th2 cell subsets. The reduction of the different subsets of T lymphocytes was associated with an intense staining with Annexin V, suggesting an apoptotic-related cell reduction. A significant decrease of Th1, Th17 and Treg cytokines and a concomitant increase of IL-4 was also observed after TCZ treatment in PBMC isolated from RA patients. CONCLUSIONS: TCZ could modify the immune imbalance in RA inducing apoptosis of Th1, Th17 and Treg cells and promoting the appearance of a Th2 response

Mehanizmi reakcija 27Al(14N, X) pri energiji snopa od 129,3 MeV

Absolute values of differential cross–sections for the formation of 6,7Li, 7,9Be, 10,11B and 12C nuclei were measured by the 14N + 27Al reaction at 129.3 MeV. The analysis of the inclusive energy spectra and angular distributions shows that the carbon and boron fragment production is well described by the projectile break–up leading to three–body final states reactions, while the lighter fragments are due both to the above–mentioned break–up and the evaporation from the 41Ca compound nucleus.Reakcijom 14N +27 Al odredili smo apsolutne vrijednosti diferencijalnih udarnih presjeka za tvorbu jezgri 6,7Li, 7,9Be, 10,11B i 12C pri energiji 129,3 MeV. Analiza inkluzivnih energijskih spektara i kutnih raspodjela pokazuje da se tvorba ugljikovih i borovih fragmenata može dobro opisati rascjepom upadne čestice koji vodi do tri tijela u konačnom stanju, dok su lakši fragmenti posljedica tog procesa i isparavanja iz složene jezgre

Mehanizmi reakcija 27Al(14N, X) pri energiji snopa od 129,3 MeV

Absolute values of differential cross–sections for the formation of 6,7Li, 7,9Be, 10,11B and 12C nuclei were measured by the 14N + 27Al reaction at 129.3 MeV. The analysis of the inclusive energy spectra and angular distributions shows that the carbon and boron fragment production is well described by the projectile break–up leading to three–body final states reactions, while the lighter fragments are due both to the above–mentioned break–up and the evaporation from the 41Ca compound nucleus.Reakcijom 14N +27 Al odredili smo apsolutne vrijednosti diferencijalnih udarnih presjeka za tvorbu jezgri 6,7Li, 7,9Be, 10,11B i 12C pri energiji 129,3 MeV. Analiza inkluzivnih energijskih spektara i kutnih raspodjela pokazuje da se tvorba ugljikovih i borovih fragmenata može dobro opisati rascjepom upadne čestice koji vodi do tri tijela u konačnom stanju, dok su lakši fragmenti posljedica tog procesa i isparavanja iz složene jezgre

A Novel White Laccase from Pleurotus ostreatus

Abstract Two laccase isoenzymes (POXA1 and POXA2) produced by Pleurotus ostreatus were purified and fully characterized. POXA1 and POXA2 are monomeric glycoproteins with 3 and 9% carbohydrate content, molecular masses of about 61 and 67 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis, of about 54 and 59 kDa by gel filtration in native conditions, and of 61 kDa by matrix-assisted laser desorption ionization mass spectrometry (only for POXA1) and pI values of 6.7 and 4.0, respectively. The N terminus and three tryptic peptides of POXA1 have been sequenced, revealing clear homology with laccases from other microorganisms, whereas POXA2 showed a blocked N terminus. The stability of POXA2 as a function of temperature was particularly low, whereas POXA1 showed remarkable high stability with respect to both pH and temperature. Both enzymes oxidize syringaldazine and ABTS (2, 2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) together with a variety of different substituted phenols and aromatic amines with the concomitant reduction of oxygen, but POXA1 is unable to oxidize guaiacol. Both enzymes were strongly inhibited by sodium azide and thioglycolic acid but not by EDTA. UV/visible absorption spectra, atomic adsorption, and polarographic data indicated the presence of 4 copper atoms/mol of POXA2 but only one copper, two zinc, and one iron atoms were found/mol of POXA1. The neutral pI and the anomalous metal content of POXA1 laccase render this enzyme unique in its structural characteristics. The lack of typical absorbance at 600 nm allows its classification as a "white" laccase

Induction and Transcriptional Regulation of Laccases in Fungi

Fungal laccases are phenol oxidases widely studied for their use in several industrial applications, including pulp bleaching in paper industry, dye decolourisation, detoxification of environmental pollutants and revalorization of wastes and wastewaters. The main difficulty in using these enzymes at industrial scale ensues from their production costs. Elucidation of the components and the mechanisms involved in regulation of laccase gene expression is crucial for increasing the productivity of native laccases in fungi. Laccase gene transcription is regulated by metal ions, various aromatic compounds related to lignin or lignin derivatives, nitrogen and carbon sources. In this manuscript, most of the published results on fungal laccase induction, as well as analyses of both the sequences and putative functions of laccase gene promoters are reviewed. Analyses of promoter sequences allow defining a correlation between the observed regulatory effects on laccase gene transcription and the presence of specific responsive elements, and postulating, in some cases, a mechanism for their functioning. Only few reports have investigated the molecular mechanisms underlying laccase regulation by different stimuli. The reported analyses suggest the existence of a complex picture of laccase regulation phenomena acting through a variety of cis acting elements. However, the general mechanisms for laccase transcriptional regulation are far from being unravelled yet

Fitting the Stocking Rate with Pastoral Resources to Manage and Preserve Mediterranean Forestlands: a Case Study

Pasture practices have affected Mediterranean forest ecosystems for millennia, and they are still quite widespread in mountainous areas. Nevertheless, in the last decades, the stability of forest ecosystems has been jeopardized due to the abandonment of traditional agro-pastoral practices, so that the gradual reduction of open areas due to progressive succession processes has caused a high increase of grazing pressure by livestock and wild ungulates feeding on forest areas. This paper aims at showing a methodological approach for evaluating the effect of applying measures in order to improve the grazing value of grasslands and ecotonal patches and lower the grazing impact on native woodlands. A protected area in Sicily (Italy) is considered as a representative case study. The analysis of remotely sensed imagery and several field surveys enabled to identify and map six different land use units subject to grazing, i.e., (1) forests; (2) grasslands (pastures dominated by palatable herbs and grasses); (3) overgrazed grasslands (dominatedby poisonous and/or thorny herbs and forbs, not palatable); (4) encroached pastures; (5) roadside firebreaks (dominated by palatable herbs) with no shrubs; and (6) wooded/ encroached roadside firebreaks. Several data were collected through sample plots selected within each land use unit, in order to assess their pastoral value. These data have been used to define current and optimal animal stock rates aiming at addressing pasture management planning towards a sustainable use of forestland and shrubland