Ecotoxicological risk of microplastics for marine organisms

La presenza di Microplastiche (MP) negli oceani rappresenta una problematica emergente per l’ecosistema marino; è ormai stato dimostrato che le MP possono essere ingerire da diverse specie di organismi, ma non è stata ancora dimostrata la capacità di trasferimento trofico e di accumulo di queste microparticelle, e sono ancora molto carenti le informazioni sui effetti avversi indotti. In questo lavoro di ricerca, il ruolo di alcuni polimeri plastici, come vettori di inquinanti chimici, è stato inizialmente valutato su campioni di macro-plastiche vergini, macro- e micro-plastiche spiaggiate. Tramite esperimenti di laboratorio, sono state invece studiate sia le cinetiche di adsorbimento del pirene e del cadmio in MP di polietilene (PE) e polistirene (PS), che la loro capacità di trasferire i composti adsorbiti ai mitili, Mytilus galloprovincialis. In questi organismi esposti sono stati analizzati diversi effetti a livello molecolare, biochimico e cellulare, incluse le risposte immunologiche, le alterazioni lisosomiali, la proliferazione perossisomiale, le difese antiossidanti, gli effetti neurotossici, genotossici oltre che il profilo di espressione genica. E’ stato inoltre ottimizzato e applicato un nuovo protocollo per l'estrazione e caratterizzazione (FT-IR) delle MP su diverse specie di pesci e invertebrati del Mar Mediterraneo. I livelli di IPA maggiori sono stati misurati nelle micro e macro-plastiche spiaggiate, mentre un efficiente assorbimento delle sostanze chimiche è stato confermato, sia per il PE che PS, con una modalità di adsorbimento tempo e dose-dipendente. Indagini istologiche hanno rivelato la presenza di MP nell’emolinfa, nelle branchie e nei tessuti digestivi dei mitili. Gli IPA adsorbiti sono stati rilasciati dalle MP e accumulati nei tessuti dei mitili che hanno mostrato alterazioni di diversi biomarker sia molecolari che cellulari. Il protocollo di estrazione sviluppato ha permesso di dimostrare la presenza di MP nello stomaco e, per la prima volta, nel fegato di cefali esposti. Studi sul campo hanno evidenziato la presenza di particelle nel 38% degli organismi selvatici analizzati, con una maggior frequenza di MP sotto forma di frammenti e linee, costituiti preferenzialmente da PE, PS e nylon. In conclusione, questa tesi fornisce nuove informazioni sui rischi ecotossicologici delle MP per gli organismi marini oltre che un’importante linea di base sul livello di contaminazione da MP nel biota Mediterraneo.Microplastics (MPs) are a growing but still unexplored environmental concern for marine organisms. Although several species can ingest MPs, a clear evidence of their accumulation pathways, trophic transfer and adverse effects is still lacking. In this thesis, the potential role of MPs as vectors of chemical pollutants was initially evaluated in virgin macroplastics, beached macro- and microplastics; laboratory experiments further characterized adsorbing kinetics of pyrene and cadmium on polyethylene (PE) and polystyrene (PS) MPs, and their capability to transfer adsorbed pyrene to mussels Mytilus galloprovincialis. In these organisms, several molecular, biochemical and cellular effects were analyzed in term of immunological responses, lysosomal alterations, peroxisomal proliferation, antioxidant and neurotoxic effects, genotoxicity and gene expression profile. A new protocol for extraction and FT-IR characterization of MPs in marine organisms was optimized and applied to several species of Mediterranean fish and invertebrates. Load of PAHs was higher in weathered and micron-sized particles, and an efficient adsorption of chemicals was confirmed with a time- and dose-dependent trend for both PE and PS. Histological analyses revealed occurrence of ingested MPs in haemolymph, gills and digestive tissues of mussels. Adsorbed PAHs were desorbed from MPs and bioavailable for mussels that showed many altered several molecular and cellular biomarkers. The developed extraction protocol allowed to demonstrate the presence of MPs in the stomach and, for the first time, in liver of exposed Mugil cephalus. Field studies highlighted the occurrence of MPs in 38% of analyzed wild organisms, mostly represented by fragments and lines, while PE, PS and nylon were the dominant polymers. In conclusion, this thesis provides new insights on the ecotoxicological risks of MPs for marine organisms and an important baseline for assessing the level of MPs contamination in Mediterranean biot

Arbuscular mycorrhizal fungi affect total phenolics content and antioxidant activity in leaves of oak leaf lettuce varieties

Plant secondary metabolites are considered key bioactive compounds for a healthy diet. Arbuscular mycorrhizal fungi (AMF) may interact with host plant metabolism, inducing the accumulation of health-promoting phytochemicals and antioxidant molecules. Lettuce is a largely consumed vegetable, which may interact with AMF to alter its content of secondary metabolites and natural antioxidants molecules, as previously shown in cultivars belonging to var. capitata or var. longifolia. In this study, the effects of red and green leaf Lactuca sativa var. crispa inoculation with different AMF species, Rhizoglomus irregulare and Funneliformis mosseae, were investigated, by assessing the total phenolics and anthocyanins content, and the antioxidant activity of leaf tissue. A significant increase of antioxidant activity and of phenolics were observed in plants of both cultivars inoculated with R. irregulare, compared to non inoculated plants. Likewise, anthocyanins (in red leaf lettuce) were more abundant in inoculated plants than in controls. Altogether, the results indicate that R. irregulare strain showed a stronger ability than F. mosseae in affecting plant metabolism and that mycorrhizal inoculation may be used to enhance concentration of phenolics in leaf type lettuces, provided that a suitable AMF is selected

Characterisation of arbuscular mycorrhizal fungi in roots by means of epifluorescence microscopy and molecular methods

In this work we characterised different species of the arbuscular mycorrhizal (AM) fungal genusGlomus by using Restriction Fragment Length Polymorphisms (RFLPs), combining epifluorescence microscopy and nested PCR, for the easy retrieval of AM fungal DNAin planta. Epifluorescence microscopy allowed us to select highly colonised root segments for DNA extraction, enhancing the chance of fungal DNA amplification by PCR. The primer pair used in nested PCR after a first reaction performed by using NS31/AM1 primers, showed a high specificity forGlomus DNA amplification and yielded enough fungal DNA for RFLP analyses. RFLP patterns of PCR products from colonised roots matched with reference profiles obtained from spores and discriminated the fourGlomus species tested. The method described may be useful for checking the establishment of mycorrhizal colonisation by the inoculated fungal species in controlled experimental conditions

Diversity of a phosphate transporter gene among species and isolates of arbuscular mycorrhizal fungi

Arbuscular mycorrhizal fungi (AMF) are a key group of beneficial obligate biotrophs, establishing a mutualistic symbiosis with the roots of most land plants. The molecular markers generally used for their characterization are mainly based on informative regions of nuclear rDNA (SSU-ITS-LSU), although protein-encoding genes have also been proposed. Within functional genes, those encoding for phosphate transporters (PT) are particularly important in AMF, given their primary ability to take up Pi from soil, and to differentially affect plant phosphate nutrition. In this work, we investigated the genetic diversity of PT1 gene sequences and sequences of the taxonomically relevant SSU-ITS-LSU region in two isolates of the species Funneliformis coronatus, three isolates of the species Funneliformis mosseae and two species of the genus Rhizoglomus, originated from geographically distant areas and cultured in vivo. Our results showed that partial PT1 sequences not only successfully differentiated AMF genera and species like ribosomal gene sequences but also highlighted intraspecific diversity among F. mosseae and F. coronatus isolates. The study of functional genes related to the uptake of key mineral nutrients for the assessment of AMF diversity represents a key step in the selection of efficient isolates to be used as inocula in sustainable agriculture

DEVELOPMENT OF YOGYACCOUNTING MONOPOLY EDUCATIVE GAME AS LEARNING MEDIA BASED COMPUTER FOR INTRODUCTION TO FINANCE AND ACCOUNTING SUBJECT AT GRADE X SMK NEGERI 1 BANTUL ACADEMIC YEAR OF 2014/2015

This research aims to: 1) developing educative game “Yogyaccounting Monopoly” as accounting learning media based computer for students at SMK Negeri 1 Bantul, 2) knowing about the feasibility of educative game “Yogyaccounting Monopoly” as accounting learning media based computer for students at SMK Negeri 1 Bantul based on validation/assessment of material experts, media experts, and practitioners, 3) knowing about the students’s response to the educative game “Yogyaccounting Monopoly” as accounting learning media based computer. This research was a Research and Development (R & D). This research conducted with development model ADDIE which consist of five development stages; 1) analysis, 2) design, 3) develop, 4) implementation, dan 5) evaluation. Validation stages was conducted with product validation by material experts of accounting introduction, learning media experts, and learning practitioner of Introduction to finance and accounting. Product was tested on 31 students of grade X AK 4 at SMKN 1 Bantul. Based on research results on the development of learning media “Yogyaccounting Monopoly” was feasible to be used in introduction to finance and accounting subject for grade X SMK 1 Bantul. It was proved from the assessment score by material experts that feasibility of material relevance aspects was 4,5; evaluation aspects was 4,5 and effects of learning strategies aspects was 5,0 so would result an average of 4,67 with the category of "Strongly Feasible". The assessment score by media experts that feasibility of language aspect was 4,0; software engineering aspect was 4,25 and visual communication was 4, so would result an average of 4,19 with the category of "Feasible". The assessment score by learning practitioners experts that feasibility of material relevance aspect was 4,0; evaluation aspect was 4,25; language aspect was 4,0 and software engineering aspect was 4,25 so would result an average of 4,13 with the category of "Feasible". Students response that feasibility of material relevance aspect was 4,36; evaluation aspect was 4,26; language aspect was 4,35; visual communication aspect was 4,04 and software engineering aspect was 4,44 so would result an average of 4,29 with the category of “ Strongly Feasible”. Key Word: Educative Game, Learning Media, Introduction to Finance and Accountin

Transición de la natalidad de Siles (Jaén) en el siglo XX : tasa bruta, proporción secundaria por sexos, generalidad, ilegitimidad y estacionalidad

En el presente trabajo se analiza la transición de la tasa bruta, proporción secundaria por sexos, gemelaridad, ilegitimidad y estacionalidad de la natalidad en la población de Siles (Jaén) a lo largo del siglo XX. Se ha fotografiado digitalmente e informatizado todas las actas de bautizos registradas desde 1900 a 1999 en los libros originales de la parroquia del pueblo, y tomado datos en el Registro Civil, totalizando 10.793 nacimientos. La tasa bruta ha pasado de 51,44‰ en 1900 a 11,51‰ en 1999. Se registra para la centuria una tasa de masculinidad del 102,65. Las frecuencias de gemelos presentan un índice total de gemeralidad de 8,98‰; el 4,447‰ en la dizigótica y el 4,54‰ en la monozigótica. La tasa global de ilegitimidad observada es del 4,52%. La estacionalidad registra máximos en el mes de agosto y mínimos en noviembre y diciembre. Se estima que la emigración ha sido el factor que más ha influido en la transición de la natalidad.