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A comparison between two recently developed, PCR-based DNA marker technologies (amplified fragment length polymorphism, AFLP; inverse sequence-lagged repeat analysis, ISTR) was carried out in a group of 19 Vitis vinifera L. accessions, including 13 putative Sangiovese-related grapevines and 6 ''coloured'' ecotypes whose fruits are of importance for conferring intense red colour to the wine. A large amount of polymorphic DNA fragments was revealed by both molecular techniques: 8 different AFLP and 5 ISTR primer combinations generated 264 and 249 polymorphic markers, respectively. Similarity relationships among the accessions were described by cluster analysis. The AFLP analysis revealed the existence of a uniform group for the Sangiovese (SG) ecotypes showing a high degree of genetic relatedness for the members of this cultivar. Among the coloured ecotypes (CLR), variability was more evident. Only the so called Colorino americano ecotype significantly diverged from both groups. ISTR analysis confirmed the genetic dissimilarity of Colorino americano and the existence of the SG and CLR groups, but in addition detected a higher proportion of polymorphism among the Sangiovese accessions compared to AFLP analysis. Sangiovese forte and Saragiolo apparently differed from the other SG-related grapevines in agreement with AFLP results. It is possible that the observed genetic dissimilarity between Sangiovese forte, Saragiolo and other SG-related types could be interpreted by the putative polyclonal origin of many grapevine cultivars, a concept which is generally accepted by the grapevine research community. Both AFLP and ISTR appear to represent innovative, efficient and sensitive molecular tools for investigating genetic diversity among Vitis vinifera ecotypes and for the eventual identification of clones., VITIS - Journal of Grapevine Research, Vol. 35 No. 4 (1996): Vitis

International audience

The European Cooperation in Science and Technology (COST) Action FP0905 is aimed at evaluating the scientific knowledge of genetically modified trees (GMT) related to biosafety protocols and coordinating existing and new information from various European countries. This will help to provide a basis for future EU policy and regulation recommendations regarding the use of GM forest trees. In this context, the Action work plan is organized in 4 Working Groups (WGs) focuses on: i) the biological characterization of GMTs aiming to evaluate existing knowledge including the experience from expert scientists in the field of forest GMTs (WG1), ii) the assessment of possible environmental impacts and monitoring of GMTs in the whole production chain from plantation to final products (WG2); iii) the socio-economic implications and public acceptance and concerns of potential use of GM forest trees and R&D investments in the framework of Cost-Benefit Analysis (WG3) and iv) increasing public awareness and understanding of GM forest plantations by providing science-based information through management of the www.cost-action-fp0905.eu dynamic website (WG4). A large, but diverse, body of knowledge on the environmental effects and biosafety issues of transgenic trees and other transgenic organisms has been acquired in many countries over approximately the past 25 years. Because of the potential unification of European states, there is now an urgent need to compile, collate, and analyze this scattered knowledge in order to create a unique platform of knowledge particular to the European environment. For this, WG1 has launched the building of a database that gather the current knowledge on field trials and greenhouse experiments with transgenic trees. The knowledge gained will ultimately be used to guide the safe use and management of GMTs in forest tree plantations and to protect forest ecosystems. To support this goal, WG2 is launching and moving through expert-based surveys to: 1) evaluate the environmental impacts of the GMTs already developed and 2) assess the efficiency of existing transgene containment strategies 3) investigate effective pre- and post-market monitoring techniques. The expected outcome from WG1 is to develop a clear factual overview of the status of GMTs in European and non-European countries in order to provide sound scientific data for risk assessment to be further evaluated and monitored in WG2. In addition, it is very important to understand the kind of policies needed to meet the concerns of the society in relation to the possible use of GM trees which are widely spread in many of the Europe countries. Therefore, two types of surveys are being conducted worldwide by WG3: one to define a set of environmental and socio-economic indicators to be included in Cost-Benefit Analyses, and a Knowledge Attitude Perception (KAP) survey to explore public attitude towards adoption of transgenic forest trees. The cross- country results of the KAP surveys are expected to provide policy support to the European Commission with regards to public acceptance of transgenic forest trees and their potential conflicts of values. On the other hand, through WG4, accurate and science-based information is being communicated through our website to help educate the general public on technical, socio-economic and environmental aspects of GM forest trees. In addition, the website provides an open discussion forum on transgenic forest biotechnology and biosafety, as well as on the potential impact of transgenic tree plantations on the current established forestry practices. Preliminary results of this research will be reported.

Il Parco Regionale di Migliarino-San Rossore-Massaciuccoli in Toscana è stato scelto come sito di studio nell'ambito del progetto europeo LIFE08 NAT/IT/342. Questo progetto è volto allo sviluppo di un indice di monitoraggio rapido (quick monitoring index, QMI) per valutare il rischio potenziale generato da piante transgeniche sugli ecosistemi agroforestali. Per questo scopo sono state selezionate tre aree di studio per determinarne la biodiversità vegetale, animale e dei microrganismi del suolo. Sono state prese particolarmente in considerazione aree vicine a superfici agricole dove potrebbero essere coltivate piante geneticamente modificate (PGM). Per ridurre il rischio di inquinamento genico di aree naturali da parte di polline di PGM, è utile sviluppare un sistema di monitoraggio efficiente che tenga conto della diversità genetica e di possibili incroci tra specie affini. Per quando riguarda la biodiversità arborea, nelle aree di studio sono state studiate specie del genere Populus. Le aree di studio selezionate per il pioppo distano tra loro 8 Km: l'area A è situata nella Tenuta di San Rossore ed è caratterizzata da un bosco misto composto da Populus alba, P. x canescens, Fraxinus angustifolia e Alnus glutinosa.; l'area B è situata presso il Lago di Massaciuccoli ed è costituita da una popolazione frammentata di Populus sp Sono state esaminate le popolazioni di Populus presenti nelle due aree insieme a due varietà coltivate. Le due aree sono state anche caratterizzate per la loro copertura erbacea. Sono stati utilizzati marcatori microsatelliti nucleari per determinare la variabilità genetica delle popolazioni di pioppo e il livello d'incrocio tra le popolazioni spontanee e quelle coltivate. Inoltre sono state studiate le popolazioni d'insetti presenti sui maschi e sulle femmine di pioppo durante il periodo di fioritura. Per determinare la densità e la composizione delle specie erbacee, nelle aree di studio è stato utilizzato l'approccio multiscala standardizzato di Dengler. Le possibili variazioni di distribuzione e/o densità delle specie sono state valutate seguendo l'approccio di Braun Blanquet. Le osservazioni fatte finora sullo strato erbaceo mostrano una certa variabilità nella distribuzione delle specie tra un quadrato e l'altro, dovuta probabilmente alle caratteristiche micro-topologiche del sito di studio. L'analisi della biodiversità degli artropodi presenti sulle infiorescenze del pioppo indica un'evidente differenza tra alberi dei due sessi e un numero piuttosto basso di specie. Nelle infiorescenze maschili predominano le larve di dittero seguite da coleotteri coccinellidi e ragni, nelle infiorescenze femminili, invece, predominano i ragni seguiti dai coleotteri curculionidi e dalle larve di lepidotteri. I dati dei microsatelliti sono stati analizzati con il software Geneland. I risultati di questa analisi mostrano tre gruppi distinti nell'area A. Il primo gruppo è composto da individui con lo stesso genotipo, probabilmente polloni, mentre gli altri due gruppi mostrano flusso genico all'interno e tra gruppi indicando che le altre specie arboree nell'area impediscono solo in parte il flusso genico. Nell'area B sono presenti due gruppi che mostrano un livello più alto di flusso genico rispetto all'area A. Questa popolazione, infatti, si trova in un'area più aperta in cui di solito il flusso genico è favorito. In generale il livello di biodiversità nelle aree è alto e in particolare il flusso genico nel lago di Massaciuccoli è considerevole. Quindi una possibile coltivazione di pioppo transgenico vicino a queste aree protette potrebbe influenzare la loro diversità. Lo sviluppo del QMI basato su dati sperimentali è in corso.

The present study investigated the extent to which physiological and biochemical traits varied because of root-zone salinity in three Mediterranean evergreens differing greatly in their strategies of salt allocation at an organismal level: the ‘salt-excluders’, Olea europaea L. and Phillyrea latifolia L. (both Oleaceae), and Pistacia lentiscus L., which, instead, largely usesNa+ and Cl_for osmotic adjustment. Both Oleaceae spp. underwent severe leaf dehydration and reduced net photosynthesis and whole-plant growth to a significantly greater degree than did P. lentiscus. Osmotic adjustment in Oleaceae mostly resulted from soluble carbohydrates, which, in turn, likely feedback regulated net photosynthesis. Salt stress reduced the actual efficiency of PSII photochemistry (FPSII) and enhanced the concentration of de-epoxided violaxanthincycle pigments in O. europaea and P. latifolia. Phenylpropanoid metabolism was upregulated by salt stress to a markedly greater degree in O. europaea and P. latifolia than in P. lentiscus. In contrast, species-specific variations in leaf lipid peroxidation were not observed in response to salinity stress. The results suggest that the species-specific ability to manage the allocation of potentially toxic ions out of sensitive leaf organs, other than affecting physiological responses, largely determined the extent to which leaf biochemistry, mostly aimed to counter salt-induced oxidative damage, varied in response to salinity stress.

Problems, such as the impact on biodiversity and the potential escapes of the trans-genes into wild populations can be amplified in the case of long-lived forest species. Therefore, studies of possible environmental impacts of transgenic forest plants (poplar) in the forest environment are important. Few studies have been published considering the alteration of the transgenic host plant genome for the possibility that transgenes can also cause pleiotropic, often undesirable alterations in plant metabolism and physiology. Other studies carried out so far, concerned the use of transgenesis to obtain poplar mutants for genomic studies or poplar breeding just for an economic purpose. The screening of the transgenic plants carrying the cry gene is in course, and the evaluation of the copy number as well as expression of the inserted gene is under evaluation. The final aim is to unravel possible pleiotropic metabolic effects in the transgenic trees following cry gene expression in P. alba and P. tremula x P. tremuloides transgenic lines.

Preliminary results of different cry transgenic poplar lines.

Constitutive expression of the rice cold-inducible Osmyb4 gene in transgenic Arabidopsis (Arabidopsis thaliana) plants improves adaptive responses to cold and drought stress, most likely due to the constitutive activation of several stress-inducible pathways and to the accumulation of several compatible solutes (e.g., glucose, fructose, sucrose, proline, glycine betaine and some aromatic compounds). Although the Osmyb4 gene seems able to activate stress responsive pathways in different species, we previously reported that its specific effect on stress tolerance depends on the transformed species. In the present work, we report the effects of the Osmyb4 expression for improving the stress response in apple (Malus pumila Mill.) plants. Namely, we found that the ectopic expression of the Myb4 transcription factor improved physiological and biochemical adaptation to cold and drought stress and modified metabolite accumulation. Based on these results it may be of interest to use Osmyb4 as a tool for improving the productivity of woody perennials under environmental stress conditions.

Problems, such as the impact on biodiversity and the potential escapes of the trans-genes into wild populations can be amplified in the case of long-lived forest species. Thus, studies of possible environmental impacts of transgenic forest plants (poplar) in the forest environment are important. Although a number of studies have been carried out so far considering the use of transgenesis for genomic studies, only little is known about biosafety issues of transgenic trees foreseen for breeding purposes. As a model transgenic tree system, we have produced Populus alba and P. tremula transgenic lines carrying the cry gene from Bacillus thuringiensis. These transgenic are under screening to unravel possible pleiotropic metabolic effects in the transgenic trees following cry gene expression. It is also not known how the use of cry-transgenic poplars affects composition and activities of soil microbial communities which affect soil health. In addition to foreign DNA, proteins from crytransgenic poplars can be released into the soil via plant residues and their persistence can constitute not only a selective pressure towards the target organisms, but also a possible threat for non-target ones with unpredictable consequences on the food chain sustained by these organisms. The work is carried out on an integrated and multidisciplinary approach (physiology, soil microbiology and biochemistry, genomic, proteomic, and microscopy), involving different scientific groups with different and specific expertise, to assess the mentioned biosafety issues important for forest trees. The screening of the transgenic plants is in course, and the evaluation of the number copy of the inserted gene as well as expression of the inserted gene is under evaluation.

Risks analyses in cry transgenic popla

A cryopreservation procedure by dehydration and direct immersion in liquid nitrogen was developed for seeds of four polyembryonic Citrus species, and the sexual or nucellar origin of the recovered seedlings was investigated. Seeds of three species could be desiccated in a sterile air flow to 16% (C. sinensis) or 10% (C. aurantium and C. limon) moisture content with a negligible reduction in germination levels. Differently, the germinability of C. deliciosa seeds dropped to 50% after drying to 15% moisture content. Following dehydration treatments, a reduction in the average number of seedlings per germinated seed was always observed. However, all four species benefited from desiccation in terms of protection during immersion in liquid nitrogen, with C. sinensis and C. aurantium showing the greatest survival (93% germination) after cryopreservation. The Inter-Simple Sequence Repeat analysis of seedlings recovered from cryopreserved seeds showed that the dehydration/cryopreservation procedure promotes the germination of zygotic embryos and reduces the number of apomictic seedlings per seed.

A cryopreservation procedure by dehydration and direct immersion in liquid nitrogen was developed for seeds of four polyembryonic Citrus species, and the sexual or nucellar origin of the recovered seedlings was investigated. Seeds of three species could be desiccated in a sterile air flow to 16% (C. sinensis) or 10% (C. aurantium and C. limon) moisture content with a negligible reduction in germination levels. Differently, the germinability of C. deliciosa seeds dropped to 50% after drying to 15% moisture content. Following dehydration treatments, a reduction in the average number of seedlings per germinated seed was always observed. However, all four species benefited from desiccation in terms of protection during immersion in liquid nitrogen, with C. sinensis and C. aurantium showing the greatest survival (93% germination) after cryopreservation. The Inter-Simple Sequence Repeat analysis of seedlings recovered from cryopreserved seeds showed that the dehydration/cryopreservation procedure promotes the germination of zygotic embryos and reduces the number of apomictic seedlings per seed.

A cryopreservation procedure by dehydration and direct immersion in liquid nitrogen was developed for seeds of four polyembryonic Citrus species, and the sexual or nucellar origin of the recovered seedlings was investigated. Seeds of three species could be desiccated in a sterile air flow to 16 percent (C. sinensis) or 10 percent (C. aurantium and C. limon) moisture content with a negligible reduction in germination levels. Differently, the germinability of C. deliciosa seeds dropped to 50 percent after drying to 15 percent moisture content. Following dehydration treatments, a reduction in the average number of seedlings per germinated seed was always observed. However, all four species benefited from desiccation in terms of protection during immersion in liquid nitrogen, with C. sinensis and C. aurantium showing the greatest survival (93 percent germination) after cryopreservation. The Inter-Simple Sequence Repeat analysis of seedlings recovered from cryopreserved seeds showed that the dehydration/cryopreservation procedure promotes the germination of zygotic embryos and reduces the number of apomictic seedlings per seed.