Genotipagem de polimorfismos associados com sistemas de macho-esterilidade em acessos de cebola adaptados ao Brasil

A produção em escala comercial de sementes híbridas de cebola (Allium cepa) tem sido conduzida com o emprego de dois sistemas de macho-esterilidade do tipo genética-citoplasmática (CMS-S e CMS-T) em associação ao citoplasma normal (macho-fértil). No entanto, a análise molecular desses diferentes tipos citoplasmáticos ainda não está disponível para um grande número de acessos de cebola adaptados para cultivo em regiões tropicais. Além de adaptação às condições edafoclimáticas do Brasil, muitos desses acessos apresentam tolerância a doenças, sendo de potencial valor como genitores de híbridos. O presente trabalho visou identificar os tipos citoplasmáticos de acessos de cebola de diferentes grupos morfoagronômicos de interesse para o melhoramento genético no Brasil, usando a reação da polimerase em cadeia (PCR) com 'primers' específicos para regiões polimórficas do genoma mitocondrial de cebola. Foi observada, nos 66 acessos amostrados, a presença dos três principais tipos de citoplasma descritos para cebola (S, N e T). Foi constatada maior frequência do citoplasma S (56%) seguido do citoplasma T (25,8%). Em 18,2% das amostras, foi encontrado exclusivamente o citoplasma N. Essa caracterização pode ser útil para guiar a escolha de materiais genéticos dentro dos programas de melhoramento com objetivo de desenvolver cultivares híbridas adaptadas às condições tropicais.

Phytoremediation of mercury by terrestrial plants

Mercury (Hg) pollution is a global environmental problem. Numerous Hg-contaminated sites exist in the world and new techniques for remediation are urgently needed. Phytoremediation, use of plants to remove pollutants from the environment or to render them harmless, is considered as an environment-friendly method to remediate contaminated soil in-situ and has been applied for some other heavy metals. Whether this approach is suitable for remediation of Hg-contaminated soil is, however, an open question. The aim of this thesis was to study the fate of Hg in terrestrial plants (particularly the high biomass producing willow, Salix spp.) and thus to clarify the potential use of plants to remediate Hg-contaminated soils. Plants used for phytoremediation of Hg must tolerate Hg. A large variation (up to 30-fold difference) was detected among the six investigated clones of willow in their sensitivity to Hg as reflected in their empirical toxicity threshold (TT95b), the maximum unit toxicity (UTmax) and EC50 levels. This gives us a possibility to select Hg-tolerant willow clones to successfully grow in Hgcontaminated soils for phytoremediation. Release of Hg into air by plants is a concern when using phytoremediation in practice. No evidence was found in this study that Hg was released to the air via shoots of willow, garden pea (Pisum sativum L. cv Faenomen), spring wheat (Triticum aestivum L. cv Dragon), sugar beet (Beta vulgaris L. cv Monohill), oil-seed rape (Brassica napus L. cv Paroll) and white clover (Trifolium repens L.). Thus, we conclude that the Hg burden to the atmosphere via phytoremediation is not increased. Phytoremediation processes are based on the ability of plant roots to accumulate Hg and to translocate it to the shoots. Willow roots were shown to be able to efficiently accumulate Hg in hydroponics, however, no variation in the ability to accumulate was found among the eight willow clones using CVAAS to analyze Hg content in plants. The majority of the Hg accumulated remained in the roots and only 0.5-0.6% of the Hg accumulation was translocated to the shoots. Similar results were found for the five common cultivated plant species mentioned above. Moreover, the accumulation of Hg in willow was higher when being cultivated in methyl-Hg solution than in inorganic Hg solution, whereas the translocation of Hg to the shoots did not differ. The low bioavailability of Hg in contaminated soil is a restricting factor for the phytoextraction of Hg. A selected tolerant willow clone was used to study whether iodide addition could increase the plant-accumulation of Hg from contaminated soil. Both pot tests and field trials were carried out. Potassium iodide (KI) addition was found to mobilize Hg in contaminated soil and thus increase the bioavailability of Hg in soils. Addition of KI (0.2–1 mM) increased the Hg concentrations up to about 5, 3 and 8 times in the leaves, branches and roots, respectively. However, too high concentrations of KI were toxic to plants. As the majority of the Hg accumulated in the roots, it might be unrealistic to use willow for phytoextraction of Hg in practice, even though iodide could enhance the phytoextraction efficiency. In order to study the effect of willow on various soil fractions of Hg-contaminated soil, a 5-step sequential soil extraction method was used. Both the largest Hg-contaminated fractions, i.e. the Hg bound to residual organic matter (53%) and sulphides (43%), and the residual fraction (2.5%), were found to remain stable during cultivations of willow. The exchangeable Hg (0.1%) and the Hg bound to humic and fulvic acids (1.1%) decreased in the rhizospheric soil, whereas the plant accumulation of Hg increased with the cultivation time. The sum of the decrease of the two Hg fractions in soils was approximately equal to the amount of the Hg accumulated in plants. Consequently, plants may be suitable for phytostabilization of aged Hg-contaminated soil, in which root systems trap the bioavailable Hg and reduce the leakage of Hg from contaminated soils.

Sintesi e proprietà di biochar da biomasse residuali

Il termine biochar definisce il prodotto solido derivante dalla pirolisi di un qualsiasi materiale organico, con lo specifico scopo di essere applicato nei suoli sia per fini agronomici che di gestione ambientale. Un suo utilizzo in maniera "responsabile" richiede però una piena comprensione delle sue proprietà e dei meccanismi che controllano la sua attività nel terreno, che dipendono dalla biomassa di partenza e dalle condizioni di sintesi tramite pirolisi. Infatti le condizioni di pirolisi, in particolare la temperatura di processo e il tempo di residenza, determinano biochar con caratteristiche differenti. In questo lavoro di tesi sono stati prodotti biochar da due diverse tipologie di biomassa residuale ampiamente disponibili (stocchi di mais e pollina). Per ciascuna biomassa sono state scelte tre condizioni di pirolisi (400°C x 20 minuti, 500°C x 10 minuti e 600°C x 5 minuti). Sui biochar ottenuti sono state effettuate le seguenti determinazioni: analisi elementare, Pirolisi‐GC‐MS, idrocarburi policiclici aromatici (IPA), acidi grassi volatili (VFA), azoto ammoniacale (N‐NH4 +), pH, conduttività elettrica e ritenzione idrica. Infine i biochar sintetizzati sono stati utilizzati per fare due test di germinazione per valutare l'effetto sulla formazione delle prime strutture di crescita delle plantule, tramite test di tossicità brevi con piastre Petri. Il primo test è stato condotto a concentrazione crescente di miscele acqua/biochar (2, 5, 40 e 100 g/L sulla base delle quantità di biochar utilizzate come ammendante nel suolo), sulla germinazione seguendo la metodologia normata dalla ISO 11269:2012. I semi utilizzati nel primo test sono stati quelli del crescione (Lepidium sativum L.) come specie dicotiledone, e del sorgo (Sorghum saccharatum M.) come monocotiledone. Il secondo saggio di tossicità eseguito è stato quello descritto dalla normativa in materia UNI 11357, valutando l'eventuale effetto di tossicità alla massima concentrazione delle varie tipologie di biochar, utilizzando come specie dicotiledoni il cetriolo (Cucumis sativus L.) ed il crescione (Lepidium sativum L.), come monocotiledone il sorgo (Sorghum saccharatum M.). Per i biochar da stocchi di mais, rappresentativi di biomasse erbacee e con diverso grado di carbonizzazione, non si osservano effetti apprezzabili alle condizioni di uso agricolo. Nel caso dei biochar da pollina si osservano invece inibizioni alla germinazione sin dalle concentrazioni più basse. In particolare, quello pirolizzato a 400°C mostra un potenziale effetto tossico più marcato, probabilmente associato ad un contenuto di IPA e VFA superiore a quello degli altri biochar.

In vitro synthesis of the light-harvesting complex into artificial membrane systems

In green plants, the function of collecting solar energy for photosynthesis is fulfilled by a series of light-harvesting complexes (LHC). The light-harvesting chlorophyll a/b protein (LHCP) is synthesized in the cytosol as a precursor (pLHCP), then imported into chloroplasts and assembled into photosynthetic thylakoid membranes. Knowledge about the regulation of the transport processes of LHCP is rather limited. Closely mimicking the in vivo situation, cell-free protein expression system is employed in this dissertation to study the reconstitution of LHCP into artificial membranes. The approach starts merely from the genetic information of the protein, so the difficult and time-consuming procedures of protein expression and purification can be avoided. The LHCP encoding gene from Pisum sativum was cloned into a cell-free compatible vector system and the protein was expressed in wheat germ extracts. Vesicles or pigment-containing vesicles were prepared with either synthetic lipid or purified plant leaf lipid to mimic cell membranes. LHCP was synthesized in wheat germ extract systems with or without supplemented lipids. The addition of either synthetic or purified plant leaf lipid was found to be beneficial to the general productivity of the expression system. The lipid membrane insertion of the LHCP was investigated by radioactive labelling, protease digestion, and centrifugation assays. The LHCP is partially protected against protease digestion; however the protection is independent from the supplemented lipids.

Development of a biocrystallisation assay for examining effects of homeopathic preparations ursing cress seedlings

A major challenge in basic research into homeopathic potentisation is to develop bioassays that yield consistent results. We evaluated the potential of a seedling-biocrystallisation method. Cress seeds (Lepidium sativum L.) germinated and grew for 4 days in vitro in Stannum metallicum 30x or water 30x in blinded and randomized assignment. 15 experiments were performed at two laboratories. CuCl2-biocrystallisation of seedlings extracted in the homeopathic preparations was performed on circular glass plates. Resulting biocrystallograms were analysed by computerized textural image analysis. All texture analysis variables analysed yielded significant results for the homeopathic treatment; thus the texture of the biocrystallograms of homeopathically treated cress exhibited specific characteristics. Two texture analysis variables yielded differences between the internal replicates, most probably due to a processing order effect. There were only minor differences between the results of the two laboratories. The biocrystallisation method seems to be a promising complementary outcome measure for plant bioassays investigating effects of homeopathic preparations.

Dithiothreitol triggers photooxidative stress and fragmentation of the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase in intact pea chloroplasts

In intact chloroplasts isolated from mature pea leaves (Pisum sativum L.), the large subunit (LSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, EC 4.1.1.39) was rapidly fragmented into several products upon illumination in the presence of 1 mM dithiothreitol (DTT). Very similar effects on LSU stability could be observed when illuminated chloroplasts were poisoned with cyanide which, like DTT, inhibits important plastid antioxidant enzymes, or when a light-dependent hydroxyl radical-producing system was added to the incubation medium. Moreover, DTT-stimulated light degradation of LSU was markedly delayed in the presence of scavengers of active oxygen species (AOS). It is therefore suggested that light degradation of LSU in the presence of DTT is mainly due to inhibition of the chloroplast antioxidant defense system and the subsequent accumulation of AOS in intact organelles. When chloroplasts were isolated from nonsenescent or senescent leaves, LSU remained very stable upon incubation without DTT, indicating that the antioxidant system was still functional in the isolated chloroplasts during leaf ageing. Our data support the notion that AOS might be important for the degradation of Rubisco in vivo under oxidative stress.

Effect of colchicine and amiprophos-methyl on the production of in vitro doubled haploid onion plants and correlation assessment between ploidy level and stomatal size

Doubled haploid onion (Allium cepa L.) plants allow the production of completely homozygous lines for a later production of hybrids. The haploid plants are normally produced using in vitro gynogenesis. The obtained haploid plantlets must be treated with different agents for doubling chromosomes. It is necessary to adjust the concentration and the length of treatment of the doubling agent. In this case, the effect of 250 and 500 mg.L-1 colchicine and 15.2; 30 and 60 mg.L- 1 amiprophos-methyl during 24 and 48 h was assessed over the rate of onion haploid plantlets chromosome doubling. The best duplication treatment was 250 mg.L-1 colchicine for 48 h, which yielded 100% of doubled haploid plants. On the other hand, a positive correlation resulted from the ploidy level and stomatal size, and a negative correlation between the level of ploidy and stomatal density. Significant differences between the stomatal length, width and density in haploid and doubled haploid plantlets were observed. An economical and quick method to test ploidy level in onion plantlets is proposed through the measurement of stomatal size and density.

Virus de la cebolla

Fil: Alcalde Lassalle, Alberto J.. Universidad Nacional de Cuyo. Facultad de Ciencias Agrarias

Pollen profiles from 8 sediment cores from the Degersee, Southern Germany

The discovery of a neolithic pile field in the shallow water near the eastern shore of the Degersee confirmed earlier palynological and sedimentological studies stating that early man was active in the region since more than 6000 years. The already available off-site data were freshly assessed, completed by additional data from old and new cores and the interpretations revised. A common time scale for the off-site data and the on-site data was obtained by AMS dating of terrestrial macro remains of the neolithic section of off-site core De_I+De_H. The ages can thus be parallelled with AMS ages of construction timber on-site. Pollen analyses from all cores provide a further time scale. The continuously and densely sampled pollen profile of the profundal zone embracing the entire Late glacial and Holocene serves as a reference. From the Boreal onwards the relative ages are transformed by AMS ages and varve counts into calibrated and absolute. A transect cored close to the neolithic pile field across the lake marl-platform demonstrates its geological architecture in the shallow water since the Lateglacial. Studies of the microfabric of thin sections of drilled cores and of box cores from the excavations demonstrate that neolithic settlements now at 2-3,5 m water depth had been erected on lake marl freshly fallen dry, thus indicating earlier lake levels dropped by 1.5-2 m. The neolithic section of the highly resolved off-site profile in the lake=s profundal zone has laminated and calcareous zones alternating with massive ones. Assemblages of diatoms and concentrations of trace elements changing simultaneously characterise the calcareous sections as deposits of low lake levels that lasted between some 40 and more than 300 years. The ages of discovered lake shore dwellings fall into calcareous segments with low lake levels. From the end of the Upper Atlantic period (F VII) appear Secondary Forest Cycles in the beech forest, a man-made sequence of repeated vegetational development with an identical pattern: With a decrease of beech pollen appear pollen of grasses, herbs and cultural indicators. These are suppressed by the light demanding hazel and birch, those again by ash, and finally by the shade demanding beech forming a new pollen peak. Seven main Forest Cycles are identified In the upper Neolithic period each comprising some 250, 450 or 800 years. They are subdivided into subcycles that can be broken down by very dense sampling in even shorter cycles of decadal length. Farming settlers have caused minor patchy clearances of the beech-mixed-forest with the use of fire. The phases of clearance coincide with peaks of charcoal and low stands of the lake levels. The Secondary Forest Cycles and the continuous occurrence of charcoal prove a continued occupation of the region. Together with the repeated restoration of the beech climax forest they point to pulsating occupation probably associated with dynamic demography. The synchronism of the many palynological, sedimentological and archaeological data point to an external forcing as the climate that affects comprehensively all these proxies. The fluctuations of the activity of the sun as manifested in the residual d14C go largely along with the proxies. The initial clearances at the begin of the forest cycles are linked to low lake levels and negative values of d14C that point to dry and warm phases of a more continental climate type. The subcycles exist independent from climatic changes, indicating that early man acted largely independent from external forces.

Pollen records and age determination of site Bokanjacko Blato

In summary, one may conclude that human influence in the Bokanjac area started in the Eneolithic or Earlier Bronze Age - the third to second millennia Cal. BC. Traces of agriculture are weak or missing in the pollen diagram but grazing is indicated. Chestnut and walnut were introduced by humans to the area in classical times. These findings are in general agreement with the results of earlier studies at coastal sites north-west and south-east of Bokanjacko Blato.

High-resolution multi-proxy record of the last glacial period from Lake Van, eastern Anatolian high plateau, Turkey

Detailed analyses of the Lake Van pollen, Ca/K ratio and stable oxygen isotope record allow the identification of millennial-scale vegetation and environmental changes in eastern Anatolia throughout the last glacial (~75-15 ka BP). The climate within the last glacial was cold and dry, with low arboreal pollen (AP) levels. The driest and coldest period corresponds to Marine Isotope Stage (MIS) 2 (~28-14.5 ka BP) dominated by the highest values of xerophytic steppe vegetation. Our high-resolution multi proxy record shows rapid expansions and contractions of tree populations that reflects variability in temperature and moisture availability. This rapid vegetation and environmental changes can be linked to the stadial-interstadial pattern of the Dansgaard-Oeschger (DO) events as recorded in the Greenland ice cores. Periods of reduced moisture availability were characterized by enhanced xerophytic species and high terrigenous input from the Lake Van catchment area. Furthermore, comparison with the marine realm reveals that the complex atmosphere-ocean interaction can be explained by the strength and position of the westerlies, which is responsible for the supply of humidity in eastern Anatolia. Influenced by diverse topography of the Lake Van catchment, larger DO interstadials (e.g. DO 19, 17-16, 14, 12 and 8) show the highest expansion of temperate species within the last glacial. However, Heinrich events (HE), characterized by highest concentrations of ice-rafted debris (IRD) in marine sediments, are identified in eastern Anatolia by AP values not lower and high steppe components not more abundant than during DO stadials. In addition, this work is a first attempt to establish a continuous microscopic charcoal record over the last glacial in the Near East, which documents an initial immediate response to millennial-scale climate and environmental variability and enables us to shed light on the history of fire activity during the last glacial.

High-resolution multi-proxy record of the last interglacial period from Lake Van, eastern Anatolian high plateau, Turkey

A high-resolution multi-proxy record from Lake Van, eastern Anatolia, derived from a lacustrine sequence cored at the 357 m deep Ahlat Ridge (AR), allows a comprehensive view of paleoclimate and environmental history in the continental Near East during the last interglacial (LI). We combined paleovegetation (pollen), stable oxygen isotope (d18Obulk) and XRF data from the same sedimentary sequence, showing distinct variations during the period from 135 to 110 ka ago leading into and out of full interglacial conditions. The last interglacial plateau, as defined by the presence of thermophilous steppe-forest communities, lasted ca. 13.5 ka, from ~129.1-115.6 ka BP. The detailed palynological sequence at Lake Van documents a vegetation succession with several climatic phases: (I) the Pistacia zone (ca. 131.2-129.1 ka BP) indicates summer dryness and mild winter conditions during the initial warming, (II) the Quercus-Ulmus zone (ca. 129.1-127.2 ka BP) occurred during warm and humid climate conditions with enhanced evaporation, (III) the Carpinus zone (ca. 127.2-124.1 ka BP) suggest increasingly cooler and wetter conditions, and (IV) the expansion of Pinus at ~124.1 ka BP marks the onset of a colder/drier environment that extended into the interval of global ice growth. Pollen data suggest migration of thermophilous trees from refugial areas at the beginning of the last interglacial. Analogous to the current interglacial, the migration documents a time lag between the onset of climatic amelioration and the establishment of an oak steppe-forest, spanning 2.1 ka. Hence, the major difference between the last interglacial compared to the current interglacial (Holocene) is the abundance of Pinus as well as the decrease of deciduous broad-leaved trees, indicating higher continentality during the last interglacial. Finally, our results demonstrate intra-interglacial variability in the low mid-latitudes and suggest a close connection with the high-frequency climate variability recorded in Greenland ice cores.

Pollen and macrofossils profiles and age determination from three cores in Northeastern Kansas, U.S.A.

Two marshes near Muscotah and Arrington, Atchison County, northeastern Kansas, yielded a pollen sequence covering the last 25,000 yrs of vegetation development. The earliest pollen spectra are comparable with surface pollen spectra from southern Saskatchewan and southeastern Manitoba and might indicate a rather open vegetation but with some pine, spruce, and birch as the most important tree species, with local stands of alder and willow. This type of vegetation changed about 23,000 yrs ago to a spruce forest, which prevailed in the region until at least 15,000 yrs ago. Because of a hiatus, the vegetation changes resulting in the spread of a mixed deciduous forest and prairie, which was present in the region from 11,000 to 9,000 yrs ago, remain unknown. Prairie vegetation, with perhaps a few trees along the valleys, covered the region until about 5,000 yrs ago, when a re-expansion of deciduous trees began in the lowlands.

Palynology on sediment profile Tso Kar in Ladakh

Palynological investigation of a 410 cm long core section from Tso Kar (33°10'N, 78°E, 4527 m a.s.l.), an alpine lake situated in the arid Ladakh area of NW India at the limit of the present-day Indian summer monsoon, was performed in order to reconstruct post-glacial regional vegetation and climate dynamics. The area was covered with alpine desert vegetation from ca. 15.2 to 14 kyr BP (1 kyr=1000 cal. years), reflecting dry and cold conditions. High influx values of long-distance transported Pinus sylvestris type pollen suggest prevailing air flow from the west and northwest. The spread of alpine meadow communities and local aquatic vegetation is a weak sign of climate amelioration after ca. 14 kyr BP. Pollen data (e.g. influx values of Pinus roxburghii type and Quercus) suggest that this was due to a strengthening of the summer monsoon and the reduced activity of westerly winds. The further spread of Artemisia and species-rich meadows occurred in response to improved moisture conditions between ca. 12.9 and 12.5 kyr BP. The subsequent change towards drier desert-steppe vegetation likely indicates more frequent westerly disturbances and associated snowfalls, which favoured the persistence of alpine meadows on edaphically moist sites. The spread of Chenopodiaceae-dominated vegetation associated with an extremely weak monsoon occurred at ca. 12.2-11.8 kyr BP during the Younger Dryas interstadial. A major increase in humidity is inferred from the development of Artemisia-dominated steppe and wet alpine meadows with Gentianaceae after the late glacial/early Holocene transition in response to the strengthening of the summer monsoon. Monsoonal influence reached maximum activity in the Tso Kar region between ca. 10.9 and 9.2 kyr BP. The subsequent development of the alpine meadow, steppe and desert-steppe vegetation points to a moderate reduction in the moisture supply, which can be linked to the weaker summer monsoon and the accompanying enhancement of the winter westerly flow from ca. 9.2 to 4.8 kyr BP. The highest water levels of Tso Kar around 8 kyr BP probably reflect combined effect of both monsoonal and westerly influence in the region. An abrupt shift towards aridity in the Tso Kar region occurred after ca. 4.8 kyr BP, as evidenced by an expansion of Chenopodiaceae-dominated desert-steppe. Low pollen influx values registered ca. 2.8-1.3 kyr BP suggest scarce vegetation cover and unfavourable growing conditions likely associated with a further weakening of the Indian Monsoon.