Global leaf and root transcriptome in response to cadmium reveals tolerance mechanisms in Arundo donax L.

The expected increase of sustainable energy demand has shifted the attention towards bioenergy crops. Due to their know tolerance against abiotic stress and relatively low nutritional requirements, they have been proposed as election crops to be cultivated in marginal lands without disturbing the part of lands employed for agricultural purposes. Arundo donax L. is a promising bioenergy crop whose behaviour under water and salt stress has been recently studied at transcriptomic levels. As the anthropogenic activities produced in the last years a worrying increase of cadmium contamination worldwide, the aim of our work was to decipher the global transcriptomic response of A. donax leaf and root in the perspective of its cultivation in contaminated soil. In our study, RNA-seq libraries yielded a total of 416 million clean reads and 10.4 Gb per sample. De novo assembly of clean reads resulted in 378,521 transcripts and 126,668 unigenes with N50 length of 1812 bp and 1555 bp, respectively. Differential gene expression analysis revealed 5,303 deregulated transcripts (3,206 up- and 2,097 down regulated) specifically observed in the Cd-treated roots compared to Cd-treated leaves. Among them, we identified genes related to "Protein biosynthesis", "Phytohormone action", "Nutrient uptake", "Cell wall organisation", "Polyamine metabolism", "Reactive oxygen species metabolism" and "Ion membrane transport". Globally, our results indicate that ethylene biosynthesis and the downstream signal cascade are strongly induced by cadmium stress. In accordance to ethylene role in the interaction with the ROS generation and scavenging machinery, the transcription of several genes (NADPH oxidase 1, superoxide dismutase, ascorbate peroxidase, different glutathione S-transferases and catalase) devoted to cope the oxidative stress is strongly activated. Several small signal peptides belonging to ROTUNDIFOLIA, CLAVATA3, and C-TERMINALLY ENCODED PEPTIDE 1 (CEP) are also among the up-regulated genes in Cd-treated roots functioning as messenger molecules from root to shoot in order to communicate the stressful status to the upper part of the plants. Finally, the main finding of our work is that genes involved in cell wall remodelling and lignification are decisively up-regulated in giant reed roots. This probably represents a mechanism to avoid cadmium uptake which strongly supports the possibility to cultivate giant cane in contaminated soils in the perspective to reserve agricultural soil for food and feed crops.

Genetic and Morpho-Agronomic Characterization of Sicilian Tetraploid Wheat Germplasm.

Cereal landraces are a very valuable resource in contemporary agriculture. A renewed focus for breeding purposes could ameliorate some negative consequences of modern agriculture and conventional breeding, such as the loss of genetic diversity. One strategy combining molecular genotyping and characterization of morpho-agronomic traits related to productivity is proposed to assess a group of tetraploid wheat landraces named Bufala, historically cultivated in the mountain areas of Sicily and characterized by adaptability in terms of cold tolerance, ability to grow in marginal soils, weed competitiveness and resistance to diseases. A total of 55 SSR molecular markers were used to detect patterns of diversity in 30 rivet and durum wheat genotypes. Furthermore, phenotyping was then conducted for 8 morpho-agronomic traits. Discriminant analysis of principal components (DAPC), STRUCTURE and phylogenetical analysis allowed to identify three groups, two of them genetically close and including both Bufala and Bufala-related rivet landraces. To the third group, old and more recent durum wheat varieties, constituting the outgroup, were assigned. Clustering was confirmed by Principal Component Analysis (PCA). Finally, a correlation analysis showed that Bufala genotypes are characterized by lower ear density, major ear length and later earing time compared with the other studied genotypes. The levels of diversity and population structure could be an important contribution to parent selection in tetraploid wheat breeding programs, as well as to germplasm conservation and management.

Transcriptional response of giant reed (Arundo donax L.) low ecotype to long-term salt stress by unigene-based RNAseq.

The giant reed is a fast growing herbaceous non-food crop considered as eligible alternative energy source to reduce the usage of fossil fuels. Tolerance of this plant to abiotic stress has been demonstrated across a range of stressful conditions, thus allowing cultivation in marginal or poorly cultivated land in order not to compromise food security and to overcome land use controversies. In this work, we de novo sequenced, assembled and analyzed the A. donax low G34 ecotype leaf transcriptome (RNAseq analysis) subjected to severe long-term salt stress (256.67 mM NaCl corresponding to 32 dS m-1 electric conductibility). In order to shed light upon the response to high salinity of this non model plant, we analyzed clusters related to salt sensory and signaling transduction, transcription factors, hormone regulation, Reactive Oxygen Species (ROS) scavenging and osmolyte biosynthesis, all of them showing different regulation compared to untreated plants. The analysis of clusters related to ethylene biosynthesis and signaling indicated that gene transcription is modulated towards the minimization of ethylene negative effects upon plant growth. Certainly, the photosynthesis is strongly affected since genes involved in Rubisco biosynthesis and assembly are down-regulated. However, a shift towards C4 photosynthesis is likely to occur as gene regulation is aimed to activate the primary CO2 fixation to PEP (phosphoenolpyruvate). The analysis of "carbon metabolism" category revealed that G34 ecotype under salt stress induces the expression of glycolysis and Krebs cycle related genes, this being consistent with the hypothesis that some sort of salt avoidance might be occurred in A. donax G34 low ecotype. By comparing our results with findings obtained with other giant reed ecotype, we identified several differences in the response to salt that are in accordance with the possibility that heritable phenotypic differences among clones of A. donax might be accumulated especially in ecotypes originating from distant geographical areas, despite their asexual reproduction modality. Additionally, 26,838 simple sequence repeat (SSR) markers were identified and validated. This SSR dataset definitely expands the marker catalogue of A. donax facilitating the genotypic characterization of this species.

Phytoremediation potential of Arundo donax (Giant Reed) in contaminated soil by heavy metals.

Soil pollution from heavy metals poses a serious risk for environment and public health. Phytoremediation is an eco-friendly and cheaper alternative compared to chemical-physical techniques. We carried out in vitro tests where three microorganisms Trichoderma harzianum, Saccharomyces cerevisiae and Wickerhamomyces anomalus were exposed to eight different heavy metals (one metal at a time) in order to evaluate resistance, growth and bioaccumulation capability for each metal (Ni, Cd, Cu, V, Zn, As, Pb, Hg). Taking into account the natural characteristics of T. harzianum, (resistance to environmental stress, resistance to pathogenic fungi, ability to establish symbiotic relationships with superior green plants) and the good bioaccumulation capacity for V, As, Cd, Hg, Pb shown after in vitro tests, it was chosen as a microorganism to be used in greenhouse tests. Controlled exposure tests were performed in greenhouse, where Arundo donax and mycorrhized Arundo donax with T. harzianum were exposed for 7 months at two different doses (L1 and L2) of a heavy metal mix, so as to assess whether the symbiotic association could improve the bioaccumulation capability of the superior green plant A. donax. Heavy metals were determined with ICP-MS. The average bioaccumulation percentage values of A. donax for L1 and L2 were, respectively: Ni (31%, 26%); Cd (35%, 50%); Cu (30%, 35%); As (19%, 27%); Pb (18%, 14%); Hg (42%, 45%); V (39%, 26%); Zn (23%, 9%). The average bioaccumulation percentage values of mycorrhized A. donax with T. harzianum for L1 and L2 were, respectively: Ni (27%, 38%); Cd (44%, 42%); Cu (36%, 29%); As (17%, 23%); Pb (37%, 54%); Hg (44%, 60%); V (16%, 20%); Zn (14%, 7%). A. donax showed the highest BAF (bioaccumulation factor) for Cd (0.50), Cu (0.35), As (0.27) and Hg (0.45) after exposure to L2; mycorrhized A. donax with T. harzianum showed the highest BAF for Ni (0.38), Cd (0.42), Pb (0.54) and Hg (0.60) after exposure to L2. A. donax showed the highest TF (translocation factor) values for Cd (0.28) and Hg (0.26) after exposition at L1 and L2 respectively; A. donax mycorrhized with T. harzianum showed the highest TF values for Cd (0.70), As (0.56), V (0.24), Pb (0.18) after exposition at L2, and Zn (0.30) after exposition at L1. Our study showed a good growth capability in contaminated soils and a good bioaccumulation capability of heavy metals, both for A. donax and mycorrhized A. donax with T. harzianum. Furthermore, for three metals (Ni, Pb and Hg) the bioaccumulation capability was improved by the symbiosis of T. harzianum with A. donax. So, these results proved the suitability both for A. donax and mycorrhized A. donax with T. harzianum for phytoremediation processes.

RNASeq analysis of giant cane reveals the leaf transcriptome dynamics under long-term salt stress.

BACKGROUND: To compensate for the lack of information about the molecular mechanism involved in Arundo donax L. response to salt stress, we de novo sequenced, assembled and analyzed the A. donax leaf transcriptome subjected to two levels of long-term salt stress (namely, S3 severe and S4 extreme). RESULTS: The picture that emerges from the identification of differentially expressed genes is consistent with a salt dose-dependent response. Hence, a deeper re-programming of the gene expression occurs in those plants grew at extreme salt level than in those subjected to severe salt stress, probably representing for them an "emergency" state. In particular, we analyzed clusters related to salt sensory and signaling, transcription factors, hormone regulation, Reactive Oxygen Species (ROS) scavenging, osmolyte biosynthesis and biomass production, all of them showing different regulation either versus untreated plants or between the two treatments. Importantly, the photosynthesis is strongly impaired in samples treated with both levels of salinity stress. However, in extreme salt conditions, a dramatic switch from C3 Calvin cycle to C4 photosynthesis is likely to occur, this probably being the more impressive finding of our work. CONCLUSIONS: Considered the distinct response to salt doses, genes either involved in severe or in extreme salt response could constitute useful markers of the physiological status of A. donax to deepen our understanding of its biology and productivity in salinized soil. Finally, many of the unigenes identified in the present study have the potential to be used for the development of A. donax varieties with improved productivity and stress tolerance, in particular the knock out of the GTL1 gene acting as negative regulator of water use efficiency has been proposed as good target for genome editing.

Targeted secondary metabolic and physico-chemical traits analysis to assess genetic variability within a germplasm collection of "long storage" tomatoes.

"Long storage" tomato is a crop traditionally cultivated in the Mediterranean area under no water supply, that recently has attracted the interest of breeders for its high tolerance to drought and as potential genetic source in breeding programs for water stress resistance. A collection of 28 genotypes of "long storage" tomato (Solanum lycopersicum L.) was studied for carotenoid and polyphenol profile and content, vitamin C, and other physico-chemical traits of fruits. Tomato carotenoids and polyphenols were identified and quantified using high-performance liquid chromatography coupled with diode array detection and electrospray-mass spectrometry (HPLC/DAD/ESI-MS); nineteen different phenolic compounds and six different carotenoids, for a total of 25 markers, have been detected, quantified and used to discriminate among the different landraces to find out which could be the best candidate for a medium-to-large scale cultivation. Different statistical approaches (ANOVA, Principal Components Analysis, Cluster Analysis) have been used for data analysis.