Characterization of the olive endophytic community in genotypes displaying a contrasting response to Xylella fastidiosa.

BACKGROUND: Endophytes mediate the interactions between plants and other microorganisms, and the functional aspects of interactions between endophytes and their host that support plant-growth promotion and tolerance to stresses signify the ecological relevance of the endosphere microbiome. In this work, we studied the bacterial and fungal endophytic communities of olive tree (Olea europaea L.) asymptomatic or low symptomatic genotypes sampled in groves heavily compromised by Xylella fastidiosa subsp. pauca, aiming to characterize microbiota in genotypes displaying differential response to the pathogen. RESULTS: The relationships between bacterial and fungal genera were analyzed both separately and together, in order to investigate the intricate correlations between the identified Operational Taxonomic Units (OTUs). Results suggested a dominant role of the fungal endophytic community compared to the bacterial one, and highlighted specific microbial taxa only associated with asymptomatic or low symptomatic genotypes. In addition, they indicated the occurrence of well-adapted genetic resources surviving after years of pathogen pressure in association with microorganisms such as Burkholderia, Quambalaria, Phaffia and Rhodotorula. CONCLUSIONS: This is the first study to overview endophytic communities associated with several putatively resistant olive genotypes in areas under high X. fastidiosa inoculum pressure. Identifying these negatively correlated genera can offer valuable insights into the potential antagonistic microbial resources and their possible development as biocontrol agents.

Transgenerational effects of chromium stress at the phenotypic and molecular level in Arabidopsis thaliana.

In this study, we describe the results obtained in a study of the transgenerational phenotypic effects of chromium (Cr) stress on the model plant species Arabidopsis thaliana. The F1 generation derived from parents grown under chronic and medium chronic stress showed significantly higher levels of the maximal effective concentration (EC50) compared with F1 plants generated from unstressed parents. Moreover, F1 plants from Cr-stressed parents showed a higher germination rate when grown in the presence of Cr. F1 plants derived from parents cultivated under chronic Cr stress displayed reduced hydrogen peroxide levels under Cr stress compared to controls. At lower Cr stress levels, F1 plants were observed to activate promptly more genes involved in Cr stress responses than F0 plants, implying a memory effect linked to transgenerational priming. At higher Cr levels, and at later stages, F1 plants modulated significantly fewer genes than F0 plants, implying a memory effect leading to Cr stress adaptation. Several bHLH transcription factors were induced by Cr stress in F1 but not in F0 plants, including bHLH100, ORG2 and ORG3. F1 plants optimized gene expression towards pathways linked to iron starvation response. A model of the transcriptional regulation of transgenerational memory to Cr stress is presented here, and could be applied for other heavy metal stresses.

Drought Stress Pre-Treatment Triggers Thermotolerance Acquisition in Durum Wheat.

Durum wheat is strongly affected by climatic constraints such as high temperatures and drought, which frequently lead to yield reduction. Damages due to high temperatures are related to plant thermotolerance, a trait determined by two components: basal and acquired thermotolerance. In this study, the effect of drought and heat stress imposed singularly or sequentially was investigated in ten durum wheat cultivars (cvs) at the physiological and molecular level. The traits analyzed were cell membrane stability, relative water content, proline content, and expression level of several genes for heat shock proteins (HSPs). Our results indicate that drought priming can induce the acquisition of thermotolerance in most cultivars already classified as able to acquire thermotolerance by heat pre-treatment. Proline accumulation was correlated to cell membrane stability, meaning that the most thermotolerant cvs were able to accumulate higher levels of proline. Acquired thermotolerance is also due to the activation of HSP gene expression; similarly, pre-treatment with water stress was able to activate HSPs expression. The results reported indicate that water stress plays an important role in inducing thermotolerance, comparable to mild heat stress pre-treatment. This is the first report on the effect of drought stress on the acquisition of thermotolerance.

Effects of Cadmium on Root Morpho-Physiology of Durum Wheat.

Durum wheat [Triticum turgidum L. subsp. durum (Desf.) Husn.] can accumulate a high level of Cd in grains with a significant variability depending on cultivars. Understanding how this toxic element is distributed in cereal tissues and grains is essential to improve the nutritional quality of cereal-based products. The main objective of this work was to investigate roots of durum wheat plants (cv. Iride) exposed to different Cd concentrations (0.5 and 5.0 µM) to identify the mechanisms involved in Cd management. Results showed that the root morphology was altered by Cd treatment both at macroscopic (increased number of tips and primary root length) and ultrastructural levels (cell membrane system damaged, cell walls thickened and enriched in suberin). On the other side, Cd was localized in vesicles and in cell walls, and the metal colocalized with the phytosiderophore nicotianamine (NA). Overall, data suggest that Cd is chelated by NA and then compartmentalized, through vesicular trafficking, in the root thickened walls reducing Cd translocation to the aerial organs of the plant.

Identifying plant genes shaping microbiota composition in the barley rhizosphere.

A prerequisite to exploiting soil microbes for sustainable crop production is the identification of the plant genes shaping microbiota composition in the rhizosphere, the interface between roots and soil. Here, we use metagenomics information as an external quantitative phenotype to map the host genetic determinants of the rhizosphere microbiota in wild and domesticated genotypes of barley, the fourth most cultivated cereal globally. We identify a small number of loci with a major effect on the composition of rhizosphere communities. One of those, designated the QRMC-3HS, emerges as a major determinant of microbiota composition. We subject soil-grown sibling lines harbouring contrasting alleles at QRMC-3HS and hosting contrasting microbiotas to comparative root RNA-seq profiling. This allows us to identify three primary candidate genes, including a Nucleotide-Binding-Leucine-Rich-Repeat (NLR) gene in a region of structural variation of the barley genome. Our results provide insights into the footprint of crop improvement on the plant's capacity of shaping rhizosphere microbes.

Diseases Caused by Xylella fastidiosa in Prunus Genus: An Overview of the Research on an Increasingly Widespread Pathogen.

Cultivated plants belonging to the genus Prunus are globally widespread and for some countries, are economically important crops; and they play a key role in the composition of a landscape. Xylella fastidiosa is a key threat to plant health, and several Prunus species are heavily stressed by this pathogen, such as almond, peach, and plum; many strain types of different subspecies can cause severe diseases. This review highlights different approaches to managing epidemic events related to X. fastidiosa in stone fruit plants. In fact, in most new European and Asian outbreaks, almond is the main and very common host and peach, plum, apricot, and cherry are widespread and profitable crops for the involved areas. Various diseases associated with stone fruit plants show different degrees of severity in relation to cultivar, although investigations are still limited. The development and selection of tolerant and resistant cultivars and the study of resistance mechanisms activated by the plant against X. fastidiosa infections seem to be the best way to find long-term solutions aimed at making affected areas recover. In addition, observations in orchards severely affected by the disease can be essential for collecting tolerant or resistant materials within the local germplasm. In areas where the bacterium is not yet present, a qualitative-quantitative study on entomofauna is also important for the timely identification of potential vectors and for developing effective control strategies.

Screening of Olive Biodiversity Defines Genotypes Potentially Resistant to Xylella fastidiosa.

The recent outbreak of the Olive Quick Decline Syndrome (OQDS), caused by Xylella fastidiosa subsp. pauca (Xf), is dramatically altering ecosystem services in the peninsula of Salento (Apulia Region, southeastern Italy). Here we report the accomplishment of several exploratory missions in the Salento area, resulting in the identification of thirty paucisymptomatic or asymptomatic plants in olive orchards severely affected by the OQDS. The genetic profiles of such putatively resistant plants (PRPs), assessed by a selection of ten simple sequence repeat (SSR) markers, were compared with those of 141 Mediterranean cultivars. Most (23) PRPs formed a genetic cluster (K1) with 22 Italian cultivars, including 'Leccino' and 'FS17', previously reported as resistant to Xf. The remaining PRPs displayed relatedness with genetically differentiated germplasm, including a cluster of Tunisian cultivars. Markedly lower colonization levels were observed in PRPs of the cluster K1 with respect to control plants. Field evaluation of four cultivars related to PRPs allowed the definition of partial resistance in the genotypes 'Frantoio' and 'Nocellara Messinese'. Some of the PRPs identified in this study might be exploited in cultivation, or as parental clones of breeding programs. In addition, our results indicate the possibility to characterize resistance to Xf in cultivars genetically related to PRPs.

Antioxidant Activity and Polyphenols Characterization of Four Monovarietal Grape Pomaces from Salento (Apulia, Italy).

The wine industry annually produces millions of tons of by-products rich in polyphenolic compounds that can be reused as secondary raw material in the food, cosmetic and pharmaceutical industries. The purpose of this work was to describe the presence of nutraceutical compounds and to evaluate the antioxidant activity of pomaces from three Apulian (South Italy, Italy) grape varieties (Negroamaro, Malvasia di Lecce and Primitivo) and to compare them with one of the most cultivated wines in Europe (Cabernet Sauvignon). The main classes of polyphenolic substances were characterized via high performance liquid chromatography/diode array detector/mass spectrometer time of flight (HPLC/DAD/TOF) and the antioxidant activity was evaluated with three different methods. The four investigated grape marcs have shown different polyphenols and antioxidant activities. Primitivo marc showed the higher antioxidant activity due to the excellent level of polyphenols, followed by the Negroamaro cultivar. In addition, marcs from traditional Apulian vines showed higher antioxidant activities than Cabernet Sauvignon because of an elevated level of active polyphenolic substances such as catechin, epicatechin, quercetin and its derivatives.

Molecular Responses to Cadmium Exposure in Two Contrasting Durum Wheat Genotypes.

Cadmium is a heavy metal that can be easily accumulated in durum wheat kernels and enter the human food chain. Two near-isogenic lines (NILs) with contrasting cadmium accumulation in grains, High-Cd or Low-Cd (H-Cd NIL and L-Cd NIL, respectively), were used to understand the Cd accumulation and transport mechanisms in durum wheat roots. Plants were cultivated in hydroponic solution, and cadmium concentrations in roots, shoots and grains were quantified. To evaluate the molecular mechanism activated in the two NILs, the transcriptomes of roots were analyzed. The observed response is complex and involves many genes and molecular mechanisms. We found that the gene sequences of two basic helix-loop-helix (bHLH) transcription factors (bHLH29 and bHLH38) differ between the two genotypes. In addition, the transporter Heavy Metal Tolerance 1 (HMT-1) is expressed only in the low-Cd genotype and many peroxidase genes are up-regulated only in the L-Cd NIL, suggesting ROS scavenging and root lignification as active responses to cadmium presence. Finally, we hypothesize that some aquaporins could enhance the Cd translocation from roots to shoots. The response to cadmium in durum wheat is therefore extremely complex and involves transcription factors, chelators, heavy metal transporters, peroxidases and aquaporins. All these new findings could help to elucidate the cadmium tolerance in wheat and address future breeding programs.

Influence of Bagging on the Development and Quality of Fruits.

Fruit quality is certainly influenced by biotic and abiotic factors, and a main quality attribute is the external appearance of the fruit. Various possible agronomical approaches are able to regulate the fruit microenvironment and, consequently, improve fruit quality and market value. Among these, fruit bagging has recently become an integral part of fruits' domestic and export markets in countries such as Japan, China, Korea Australia and the USA because it is a safe and eco-friendly technique to protect fruits from multiple stresses, preserving or improving the overall quality. Despite increasing global importance, the development of suitable bagging materials and, above all, their use in the field is quite laborious, so that serious efforts are required to enhance and standardize bagging material according to the need of the crops/fruits. This review provides information about the effects of bagging technique on the fruit aspect and texture, which are the main determinants of consumer choice.

Secondary Metabolites in Xylella fastidiosa-Plant Interaction.

During their evolutionary history, plants have evolved the ability to synthesize and accumulate small molecules known as secondary metabolites. These compounds are not essential in the primary cell functions but play a significant role in the plants' adaptation to environmental changes and in overcoming stress. Their high concentrations may contribute to the resistance of the plants to the bacterium Xylella fastidiosa, which has recently re-emerged as a plant pathogen of global importance. Although it is established in several areas globally and is considered one of the most dangerous plant pathogens, no cure has been developed due to the lack of effective bactericides and the difficulties in accessing the xylem vessels where the pathogen grows and produces cell aggregates and biofilm. This review highlights the role of secondary metabolites in the defense of the main economic hosts of X. fastidiosa and identifies how knowledge about biosynthetic pathways could improve our understanding of disease resistance. In addition, current developments in non-invasive techniques and strategies of combining molecular and physiological techniques are examined, in an attempt to identify new metabolic engineering options for plant defense.

Editorial for Special Issue "Heavy Metals Accumulation, Toxicity, and Detoxification in Plants".

"Heavy metals" is a collective term widely applied for the group of metals and metalloids with an atomic density above 4 g/cm3 [...].

Biochemical Changes in Leaves of Vitis vinifera cv. Sangiovese Infected by Bois Noir Phytoplasma.

Bois noir is a disease associated with the presence of phytoplasma 'Candidatus Phytoplasma solani' belonging to the Stolbur group (subgroup 16SrXII-A), which has a heavy economic impact on grapevines. This study focused on the changes induced by phytoplasma in terms of the profile and amount of secondary metabolites synthesized in the phenylpropanoid pathway in leaves of Vitis vinifera L. red-berried cultivar Sangiovese. Metabolic alterations were assessed according to the disease progression through measurements of soluble sugars, chlorophyll, and phenolic compounds produced by plant hosts, in response to disease on symptomatic and asymptomatic Bois noir-positive plants. Significant differences were revealed in the amount of soluble sugars, chlorophyll, and accumulation/reduction of some compounds synthesized in the phenylpropanoid pathway of Bois noir-positive and negative grapevine leaves. Our results showed a marked increase in phenolic and flavonoid production and a parallel decrease in lignin content in Bois noir-positive compared to negative leaves. Interestingly, some parameters (chlorophyll a, soluble sugars, total phenolic or flavonoids content, proanthocyanidins, quercetin) differed between Bois noir-positive and negative leaves regardless of symptoms, indicating measurable biochemical changes in asymptomatic leaves. Our grapevine cultivar Sangiovese results highlighted an extensive modulation of the phenylpropanoid biosynthetic pathway as a defense mechanism activated by the host plant in response to Bois noir disease.

Combined Effect of Cadmium and Lead on Durum Wheat.

Cadmium (Cd) and lead (Pb) are two toxic heavy metals (HMs) whose presence in soil is generally low. However, industrial and agricultural activities in recent years have significantly raised their levels, causing progressive accumulations in plant edible tissues, and stimulating research in this field. Studies on toxic metals are commonly focused on a single metal, but toxic metals occur simultaneously. The understanding of the mechanisms of interaction between HMs during uptake is important to design agronomic or genetic strategies to limit contamination of crops. To study the single and combined effect of Cd and Pb on durum wheat, a hydroponic experiment was established to examine the accumulation of the two HMs. Moreover, the molecular mechanisms activated in the roots were investigated paying attention to transcription factors (bHLH family), heavy metal transporters and genes involved in the biosynthesis of metal chelators (nicotianamine and mugineic acid). Cd and Pb are accumulated following different molecular strategies by durum wheat plants, even if the two metals interact with each other influencing their respective uptake and translocation. Finally, we demonstrated that some genes (bHLH 29, YSL2, ZIF1, ZIFL1, ZIFL2, NAS2 and NAAT) were induced in the durum wheat roots only in response to Cd.

Molecular Effects of Xylella fastidiosa and Drought Combined Stress in Olive Trees.

Due to global climate change, complex combinations of stresses are expected to occur, among which the interaction between pathogens and drought stress may have a significant effect on growth and yield. In this study, the Xylella fastidiosa (Xf)-resistant cultivar Leccino and the susceptible one Cellina di Nardò were subjected to (a) individual drought stress, (b) Xf infection and (c) combination of both stress conditions. Here we report the physiological response to stresses in water content in leaves and the modulation in the expression level of seven genes responsive to plant water status and pathogen infection. In Xf-resistant plants, higher expression levels are reported for genes belonging to ROS-scavenging systems and for genes involved in pathogen stress (pathogenesis-related, PR, and leucine-rich repeat genes, LRR-RLK). However, PR and LRR-RLK were not further induced by water deficit. Interestingly, the genes related to drought response (aquaporin, PIP2.1, dehydration responsive element binding, DREB, and dehydrin, DHN), which induction was higher in Cellina di Nardò compared to Leccino during drought stress, was poorly induced in Xf-susceptible plants when Xf occur. Conversely, DHN was induced by Xf presence in Leccino. These results were consistent with observations on water content. Indeed, response was similar in Leccino regardless kind of stress or combination, whereas a strong reduction was observed in Xf-susceptible plants infected by Xf or in presence of combined stresses. Thus, the reported findings indicate that resistance of Leccino to Xf could be linked to its lower resistance to water stress, probably leading to the activation of alternative defense pathways that support the plant in Xf response.

Nutraceutical Properties of Mulberries Grown in Southern Italy (Apulia).

In this work, for the first time, were analyzed mulberry genotypes grown in Apulia (Southern Italy, Salento region) were analyzed. Two local varieties of Morus alba (cv. Legittimo nero and cv. Nello) and one of Morus nigra were characterized for content in simple sugars, organic acids, phenols, anthocyanins; fruit antioxidant activity (AA) was also evaluated by three different methods (2,2-Diphenyl-1-picrylhydrazyl, DPPH; 2,2'-Azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ABTS; and Ferric reducing antioxidant potential, FRAP test). The results showed that the sugars amount ranged between 6.29 and 7.66 g/100 g fresh weight (FW) while the malic and citric acids content was low, at about 0.1-1 g/100 g FW. Mulberries are a good source of phenols which are present in higher values in M. nigra and M. alba cv. Legittimo nero (485 and 424 mg Gallic Acid Equivalent (GAE)/ 100 g FW, respectively). The high performance liquid chromatography/diode array detector/mass spectrometry (HPLC/DAD/MS) analysis identified 5 main anthocyanin compounds present in different concentrations in each variety of mulberry: cyanidin 3-sophoroside, cyanidin 3-glucoside, cyanidin 3-rutinoside, pelargonidin 3-glucoside, pelargonidin 3-rutinoside. The highest concentration of anthocyanins was determined in Morus alba Legittimo (about 300 mg/100 g FW) while the lowest content (about 25 mg/100 g FW) was measured in M. alba cv. Nello. Morus nigra showed a good AA in comparison with the different M. alba genotypes with all the used methods; its AA was equal to 33, 26 and 21 µmols Trolox/g FW when using DPPH, ABTS and FRAP tests, respectively. All genotypes showed an anti-inflammatory activity (measured by cyclooxygenase (COX) inhibitory assay) which was also compared with two commercial anti-inflammatory drugs. The data obtained support the high biological qualities of mulberry fruits and their diffusion in human nutrition.

Xylem cavitation susceptibility and refilling mechanisms in olive trees infected by Xylella fastidiosa.

In olive trees, Xylella fastidiosa colonizes xylem vessels and compromises water transport causing the olive quick decline syndrome (OQDS). The loss of hydraulic conductivity could be attributed to vessel occlusions induced both by the bacteria biofilm and by plant responses (tyloses, gums, etc.) that could trigger embolism. The ability of the infected plants to detect embolism and to respond, by activating mechanisms to restore the hydraulic conductivity, can influence the severity of the disease symptomatology. In order to investigate these mechanisms in the X. fastidiosa-resistant olive cultivar Leccino and in the susceptible Cellina di Nardò, sections of healthy olive stems were analysed by laser scanning microscope to calculate the cavitation vulnerability index. Findings indicated that the cultivar Leccino seems to be constitutively less susceptible to cavitation than the susceptible one. Among the vascular refilling mechanisms, starch hydrolysis is a well-known strategy to refill xylem vessels that suffered cavitation and it is characterized by a dense accumulation of starch grains in the xylem parenchima; SEM-EDX analysis of stem cross-sections of infected plants revealed an aggregation of starch grains in the Leccino xylem vessels. These observations could indicate that this cultivar, as well as being anatomically less susceptible to cavitation, it also could be able to activate more efficient refilling mechanisms, restoring vessel's hydraulic conductivity. In order to verify this hypothesis, we analysed the expression levels of some genes belonging to families involved in embolism sensing and refilling mechanisms: aquaporins, sucrose transporters, carbohydrate metabolism and enzymes related to starch breakdown, alpha and beta-amylase. The obtained genes expression patterns suggested that the infected plants of the cultivar Leccino strongly modulates the genes involved in embolism sensing and refilling.

Phenolic Profile and Antioxidant Activity of Italian Monovarietal Extra Virgin Olive Oils.

In the last years, the interest in Italian monovarietal oils has increased due to their specific organoleptic qualities. Extra virgin olive oils (EVOOs) are rich in phenolic compounds, secondary metabolites well known and studied for their nutraceutical properties. However, among EVOOs, there is great variability in phenolic composition due to the origin, the production technique, and mainly, the genotype. The aim of this work was to evaluate the different phenolic profiles and the antioxidant activities of monovarietal oils. The results confirm this variability. In fact, the overall content of oleuropein varies up to four times between the different genotypes (from 33.80 to 152.32 mg/kg oil), while the oleocanthal content is significant only in two oils. The antioxidant activity, determined with 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and oxygen radical absorbance capacity (ORAC) assays, is correlated with the content of total phenolic substances, with half maximal inhibitory concentration (IC50) values for the DPPH test ranging from 160 to 91 mg of oil, while the ORAC test shows values between 5.45 and 8.03 µmol Trolox equivalent (TE)/g oil.

Evaluation of Phytochemical and Antioxidant Properties of 15 Italian Olea europaea L. Cultivar Leaves.

Olive leaf extracts are of special interest due to their proven therapeutic effects. However, they are still considered a by-product of the table olive and the oil industries. In order to learn possible ways of exploiting this waste for health purposes, we investigated the phytochemical profiles and antioxidant activities in the leaves of 15 Italian Olea europaea L. cultivars grown in the same pedoclimatic conditions. The phenolic profiles and amounts of their seven representative compounds were analyzed using HPLC ESI/MS-TOF. The antioxidant activities were determined using three different antioxidant assays (DPPH, ORAC, and superoxide anion scavenging assay). Wide ranges of total phenolic content (11.39-48.62 g GAE kg-1 dry weight) and antioxidant activities (DPPH values: 8.67-29.89 µmol TE mg-1 dry weight, ORAC values: 0.81-4.25 µmol TE mg-1 dry weight, superoxide anion scavenging activity values: 27.66-48.92 µmol TE mg-1 dry weight) were found in the cultivars. In particular, the cultivars Itrana, Apollo, and Maurino, showed a high amount of total phenols and antioxidant activity, and therefore represent a suitable natural source of biological compounds for use in terms of health benefits.

Antioxidant Activity and Anthocyanin Contents in Olives (cv Cellina di Nardò) during Ripening and after Fermentation.

The olive tree "Cellina di Nardò" (CdN) is one of the most widespread cultivars in Southern Italy, mainly grown in the Provinces of Lecce, Taranto, and Brindisi over a total of about 60,000 hectares. Although this cultivar is mainly used for oil production, the drupes are also suitable and potentially marketable as table olives. When used for this purpose, olives are harvested after complete maturation, which gives to them a naturally black color due to anthocyanin accumulation. This survey reports for the first time on the total phenolic content (TPC), anthocyanin characterization, and antioxidant activity of CdN olive fruits during ripening and after fermentation. The antioxidant activity (AA) was determined using three different methods. Data showed that TPC increased during maturation, reaching values two times higher in completely ripened olives. Anthocyanins were found only in mature olives and the concentrations reached up to 5.3 g/kg dry weight. AA was determined for the four ripening stages, and was particularly high in the totally black olive fruit, in accordance with TPC and anthocyanin amounts. Moreover, the CdN olives showed a higher TPC and a greater AA compared to other black table olives produced by cultivars commonly grown for this purpose. These data demonstrate the great potential of black table CdN olives, a product that combines exceptional organoleptic properties with a remarkable antioxidant capacity.