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Epigenetic modifications play a vital role in the preservation of genome integrity and in the regulation of gene expression. DNA methylation, one of the key mechanisms of epigenetic control, impacts growth, development, stress response and adaptability of all organisms, including plants. The detection of DNA methylation marks is crucial for understanding the mechanisms underlying these processes and for developing strategies to improve productivity and stress resistance of crop plants. There are different methods for detecting plant DNA methylation, such as bisulfite sequencing, methylation-sensitive amplified polymorphism, genome-wide DNA methylation analysis, methylated DNA immunoprecipitation sequencing, reduced representation bisulfite sequencing, MS and immuno-based techniques. These profiling approaches vary in many aspects, including DNA input, resolution, genomic region coverage, and bioinformatics analysis. Selecting an appropriate methylation screening approach requires an understanding of all these techniques. This review provides an overview of DNA methylation profiling methods in crop plants, along with comparisons of the efficacy of these techniques between model and crop plants. The strengths and limitations of each methodological approach are outlined, and the importance of considering both technical and biological factors are highlighted. Additionally, methods for modulating DNA methylation in model and crop species are presented. Overall, this review will assist scientists in making informed decisions when selecting an appropriate DNA methylation profiling method.

The salinization of soil is the process of progressive accumulation of salts such as sulfates, sodium, or chlorides into the soil. The increased level of salt has significant effects on glycophyte plants, such as rice, maize, and wheat, which are staple foods for the world's population. Consequently, it is important to develop biotechnologies that improve crops and clean up the soil. Among other remediation methods, there is an environmentally friendly approach to ameliorate the cultivation of glycophyte plants in saline soil, namely, the use of microorganisms tolerant to salt with growth-promoting features. Plant growth-promoting rhizobacteria (PGPR) can improve plant growth by colonizing their roots and playing a vital role in helping plants to establish and grow in nutrient-deficient conditions. Our research aimed to test in vivo halotolerant PGPR, isolated and characterized in vitro in a previous study conducted in our laboratory, inoculating them on maize seedlings to improve their growth in the presence of sodium chloride. The bacterial inoculation was performed using the seed-coating method, and the produced effects were evaluated by morphometric analysis, quantization of ion contents (sodium, potassium), produced biomass, both for epigeal (shoot) and hypogeal (root) organs, and by measuring salt-induced oxidative damage. The results showed an increase in biomass and sodium tolerance and even a reduction of oxidative stress in seedlings pretreated with a PGPR bacterial consortium (Staphylococcus succinus + Bacillus stratosphericus) over the control. Moreover, we observed that salt reduces growth and alters root system traits of maize seedlings, while bacterial treatment improves plant growth and partially restores the root architecture system in saline stress conditions. Therefore, the PGPR seed-coating or seedling treatment could be an effective strategy to enhance sustainable agriculture in saline soils due to the protection of the plants from their inhibitory effect.

Soil salinity, due to natural phenomena or human activities, alters the water potential, which in turn affects plant growth, negatively influencing their nutrient and water uptake. Plant-growth-promoting rhizobacteria (PGPR) can be used to counteract these negative effects, especially in glycophytes. The aim of our study was to characterize physiologically, genetically, and biochemically the novel halotolerant/halophilic bacteria isolated in our previous work. We evaluated the plant-growth-promoting (PGP) features and NaCl regulation’s roles in them. In this regard, analysis based on 16S rDNA sequences confirmed that our isolates are distinct bacterial strains, probably belonging to new species, which we named Bacillus sp. M21 and M23 and Halomonas sp. QH23 and QH24. In literature, it is known that many species of Halomonas and Bacillus genera produce factors regulating plant development, such as indole-3-acetic acid (IAA), ammonium, and siderophores; and their efficiency in promoting plant growth and productivity was also demonstrated in vivo. We demonstrated that the newly isolated strains exhibit different PGP activities, highlighting how the latter are regulated by NaCl and in a strain-dependent manner. In particular, the main results showed that NaCl negatively affects the production of IAA in QH23, M21 and M23, whereas it promotes it in QH24, where it is strictly salt-dependent. Both Halomonas strains produce ammonium only in the presence of NaCl, directly correlated to its concentration. The opposite occurs in Bacillus strains, where salt reduces its production up to ten times. Overall, the results underline how halotolerance is a preferable, but not sufficient, condition for considering a PGPR potentially useful in applications aimed at improving the tolerance and productivity of crops in saline soils.

Crop adaptation to climate change is in a part attributed to epigenetic mechanisms which are related to response to abiotic and biotic stresses. Although recent studies increased our knowledge on the nature of these mechanisms, epigenetics remains under-investigated and still poorly understood in many, especially non-model, plants, Epigenetic modifications are traditionally divided into two main groups, DNA methylation and histone modifications that lead to chromatin remodeling and the regulation of genome functioning. In this review, we outline the most recent and interesting findings on crop epigenetic responses to the environmental cues that are most relevant to climate change. In addition, we discuss a speculative point of view, in which we try to decipher the “epigenetic alphabet” that underlies crop adaptation mechanisms to climate change. The understanding of these mechanisms will pave the way to new strategies to design and implement the next generation of cultivars with a broad range of tolerance/resistance to stresses as well as balanced agronomic traits, with a limited loss of (epi)genetic variability.

A high-quality transcriptome is required to advance numerous bioinformatics workflows. Nevertheless, the effectuality of tools for de novo assembly and real precision assembled transcriptomes looks somewhat unexplored, particularly for non-model organisms with complicated (very long, heterozygous, polyploid) genomes. To disclose the performance of various transcriptome assembly programs, this study built 11 single assemblies and analyzed their performance on some significant reference-free and reference-based criteria. As well as to reconfirm the outputs of benchmarks, 55 BLAST were performed and compared using 11 constructed transcriptomes. Concisely, normalized benchmarking demonstrated that Velvet–Oases suffer from the worst results, while the EvidentialGene strategy can provide the most comprehensive and accurate transcriptome of Lilium ledebourii (Baker) Boiss. The BLAST results also confirmed the superiority of EvidentialGene, so it could capture even up to 59% more (than Velvet–Oases) unique gene hits. To promote assembly optimization, with the help of normalized benchmarking, PCA and AHC, it is emphasized that each metric can only provide part of the transcriptome status, and one should never settle for just a few evaluation criteria. This study supplies a framework for benchmarking and optimizing the efficiency of assembly approaches to analyze RNA-Seq data and reveals that selecting an inefficient assembly strategy might result in less identification of unique gene hits.

Until recently, many phytoremediation studies were focused solely on a plants ability to reclaim heavy metal (HM) polluted soil through a range of different processes, such as phytoextraction and phytostabilization. However, the interaction between plants and their own rhizosphere microbiome represents a new research frontier for phytoremediation. Our hypothesis is that rhizomicrobiome might play a key role in plant wellness and in the response to external stimuli; therefore, this study aimed to shed light the rhizomicrobiome dynamics after an organic amendment (e.g., compost) and/or HM pollution (e.g., Zn), and its relation with plant reclamation ability. To reach this goal we set up a greenhouse experiment cultivating in pot an elite black poplar clone (N12) selected in the past for its excellent ability to reclaim heavy metals. N12 saplings were grown on a soil amended with compost and/or spiked with high Zn doses. At the end of the experiment, we observed that the compost amendment strongly increased the foliar size but did not affect significantly the Zn accumulation in plant. Furthermore, the rhizomicrobiome communities (bacteria and fungi), investigated through NGS, highlighted how α diversity increased in all treatments compared to the untreated N12 saplings. Soil compost amendment, as well as Zn pollution, strongly modified the bacterial rhizomicrobiome structure. Conversely, the variation of the fungal rhizomicrobiome was only marginally affected by soil Zn addition, and only partially impaired by compost. Nevertheless, substantial alterations of the fungal community were due to both compost and Zn. Together, our experimental results revealed that organic amendment increased the bacterial resistance to external stimuli whilst, in the case of fungi, the amendment made the fungi microbiome more susceptible. Finally, the greater microbiome biodiversity does not imply, in this case, a better plant wellness or phytoremediation ability, although the microbiome plays a role in the external stimuli response supporting plant life.

Hawthorn (Crataegus monogyna Jacq.) is a wild edible fruit tree of the genus Crataegus, one of the most interesting genera of the Rosaceae family. This review is the first to consider, all together, the pharmaceutical, phytochemical, functional and therapeutic properties of C. monogyna based on numerous valuable secondary metabolites, including flavonoids, vitamin C, glycoside, anthocyanin, saponin, tannin and antioxidants. Previous reviews dealt with the properties of all species of the entire genera. We highlight the multi-therapeutic role that C. monogyna extracts could have in the treatment of different chronic and degenerative diseases, mainly focusing on flavonoids. In the first part of this comprehensive review, we describe the main botanical characteristics and summarize the studies which have been performed on the morphological and genetic characterization of the C. monogyna germplasm. In the second part, the key metabolites and their nutritional and pharmaceutical properties are described. This work could be an essential resource for promoting future therapeutic formulations based on this natural and potent bioactive plant extract.

Evaluating the relative weight of the choice of cultivar and soil fertilization on potentially toxic elements (PTEs) accumulation is crucial in promoting informed decisions in the framework of regenerative agriculture. To this end, 11 PTEs (Cd, Cr, Cu, Fe, K, Mg, Mn, Na, Ni, Pb, Zn) were determined in both leaves and roots of six cultivars (Stylist, Xanadu, Aljeva, Bacio, Analena, Vincenzo) of lettuce (Lactuca sativa L.) grown side by side on mineral fertilized or biowaste compost amended soils, under greenhouse conditions. The use of multivariate and linear modelling approaches indicated that the organ and cultivar primarily account for the variability in PTE concentrations. In terms of PTE partitioning between organs, Cd and Mg were preferentially accumulated in leaves, whereas Cu, Pb, K and Zn in roots. As for the cultivar, Xanadu showed the highest concentrations of several PTEs, with Cd reaching concerning levels. Fertilization had a detectable contribution only on Cd accumulation, slightly increased in leaves by compost. Findings highlight the key role of cultivar choice in guaranteeing food safety and grant the possibility to adopt biowaste compost in regenerative agriculture without concerns about PTE accumulation enhancements in lettuce, but demand a cautionary approach in the case of Cd.

Among the most widespread plant species with clonal reproduction Arundo donax L. represents one of most studied one characterized by very low genetic biodiversity. Although it is a perennial rhizomatous tall grass native to eastern and southern Asia, it spreads only asexually in the invaded range all over the world thriving very well in a large array of pedo-climatic conditions. This ability to morphologically or physiologically adapt to a broad array of conditions could be attributed to epigenetic mechanisms. To shade light on this relevant issue, 96 stems of A. donax from spontaneous populations distributed across the Italian invaded range (island of Sardinia, Northern and Southern Italy) were analysed. Leaf DNAs were extracted and processed through AFLPs and MSAPs for defining either genetic and epigenetic profiles. Both analyses clearly showed that the A. donax populations of Sardinia island are genetically distinct from those of Italian mainland; AFLPs showed an extremely low genetic biodiversity due to vegetative reproduction, whilst, epi-biodiversity, estimated through MSAP marker, increased within the analyzed populations. These results suggest that the capability of A. donax to invade and thrive in diverse environmental conditions can be, at least, partially attributed to a higher epigenetic variability. Therefore, the different DNA methylation status may have significant and important biological meaning, in particular, in the case of invasive clonal plants such as A. donax, also for the biodiversity definition, and MSAP marker can be considered an useful and cost effective marker to reveal it.

Italy displays a high level of agrobiodiversity due to its diversified pedoclimatic zones. The Administrative Region of Campania includes several and divergent biomes, occurring close to each other. In fact, the distance between a sea level environment and that of high mountains can be less than 20 km. These environmental conditions allow the cultivation of many different crops and vegetables, represented by diverse ecotypes and varieties that are well adapted to the distribution range where they have been selected and grown. Efforts to maintain and further increase biodiversity in farming systems require a better understanding of the existing diversity created by traditional farming practices. The aim of our study was to identify and molecularly characterize several ecotypes belonging to five horticultural species commonly cultivated in Campania. In particular, we analysed five ecotypes of maize, two of garlic, four of onion, one of escarole, and two of courgette by means of simple sequence repeat (SSR) markers in order to evaluate their level of genetic biodiversity. The results reveal, for the first time, the high genetic biodiversity of horticultural ecotypes of the Campania Region. This feature is very important to improve the quality and productivity of agroecosystems.

Soil salinization and desertification due to climate change are the most relevant challenges for the agriculture of the 21st century. Soil compost amendment and plant growth promoting rhizobacteria (PGP-R) are valuable tools to mitigate salinization and desertification impacts on agricultural soils. Selection of novel halo/thermo-tolerant bacteria from the rhizosphere of glicophytes and halophytes, grown on soil compost amended and watered with 150/300 mM NaCl, was the main objective of our study. Beneficial effects on the biomass, well-being and resilience, exerted on the assayed crops (maize, tomato, sunflower and quinoa), were clearly observable when soils were amended with 20% compost despite the very high soil electric conductivity (EC). Soil compost amendment not only was able to increase crop growth and biomass, but also their resilience to the stress caused by very high soil EC (up to 20 dS m−1). Moreover, compost amendment has proved itself a valuable source of highly halo-(4.0 M NaCl)/thermo tolerant rhizobacteria (55 °C), showing typical PGP features. Among the 13 rhizobacterial isolates, molecularly and biochemically characterized, two bacterial strains showed several biochemical PGP features. The use of compost is growing all around the world reducing considerably for farmers soil fertilization costs. In fact, only in Italy its utilization has ensured, in the last years, a saving of 650 million euro for the farmers, without taking into account the environment and human health benefits. Furthermore, the isolation of halo/thermo-tolerant PGPR strains and their use will allow the recovery and cultivation of hundreds of thousands of hectares of saline and arid soils now unproductive, making agriculture more respectful of agro-ecosystems also in view of upcoming climate change.

In the absence of genetic diversity, plants rely on the capacity of phenotypic plasticity to cope with shifts in environmental conditions. Understanding the mechanisms behind phenotypic plasticity and how local phenotypic adjustments are transferred to clonal offspring, will provide insight into its ecological and evolutionary significance. Epigenetic changes have recently been proposed to play a crucial role in rapid environmental adaptation. While the contribution of epigenetic changes to phenotypic plasticity has been extensively studied in sexual reproducing model organisms, little work has been done on vegetative generations of asexual reproducing plant species. We studied the variability of DNA methylation and bud set phenology of the Lombardy poplar (Populus nigra cv. Italica Duroi), a cultivated tree representing a single genotype worldwide distributed since the eighteenth century. Bud set observations and CpG methyl polymorphisms were studied on vegetative offspring resulting from cuttings grown for one season in a common glasshouse environment. The cuttings were collected from 60 adult Lombardy poplars growing in different environments. The physiological condition of the cuttings was determined by measuring weight and nutrient condition. Methylation sensitive amplified polymorphisms were used to obtain global patterns of DNA methylation. Using logistic regression models, we investigated correlations among epigenotype, bud phenology, and the climate at the home site of the donor trees, while accounting for physiological effects. We found significant epigenetic variation as well as significant variation in bud phenology, in the absence of genetic variation. Remarkably, phenology of bud set observed at the end of the growing season in the common environment was significantly correlated with climate variables at the home site of the mother trees, specifically the average temperature of January and monthly potential evapotranspiration. Although we could not directly detect significant effects of epigenetic variation on phenology, our results suggest that, in the Lombardy poplar, epigenetic marks contribute to the variation of phenotypic response that can be transferred onto asexually reproduced offspring resulting in locally adapted ecotypes. This contributes to the growing evidence that epigenetic-based transgenerational inheritance might be relevant for adaptation and evolution in contrasting or rapidly changing environments.

Morphometric, biochemical and genetic analyses were conducted on Olea europaea L. of Campania, an area of Southern Italy highly suited to the cultivation of olive trees and the production of extra virgin olive oil (EVOO). We aimed to characterize the distribution of morphological, biochemical and genetic diversity in this area and to develop a practical tool to aid traceability of oils. Phenotypes were characterized using morphometric data of drupes and leaves; biochemical and genetic diversity were assessed on the basis of the fatty acid composition of the EVOOs and with microsatellite markers, respectively. We provide an open-source tool as a novel R package titled ‘OliveR’, useful in performing multivariate data analysis using a point and click interactive approach. These analyses highlight a clear correlation among the morphological, biochemical and genetic profiles of samples with four collection sites, and confirm that Southern Italy represents a wide reservoir of phenotypic and genetic variability.

The widespread poplar populations of Sardinia are vegetatively propagated and live in different natural environments forming large monoclonal stands. The main goals of the present study were: i) to investigate/measure the epigenetic diversity of the poplar populations by determining their DNA methylation status; ii) to assess if and how methylation status influences population clustering; iii) to shed light on the changes that occur in the epigenome of ramets of the same poplar clone. To these purposes, 83 white poplar trees were sampled at different locations on the island of Sardinia. Methylation sensitive amplified polymorphism analysis was carried out on the genomic DNA extracted from leaves at the same juvenile stage. The study showed that the genetic biodiversity of poplars is quite limited but it is counterbalanced by epigenetic inter-population molecular variability. The comparison between MspI and HpaII DNA fragmentation profiles revealed that environmental conditions strongly influence hemi-methylation of the inner cytosine. The variable epigenetic status of Sardinian white poplars revealed a decreased number of population clusters. Landscape genetics analyses clearly demonstrated that ramets of the same clone were differentially methylated in relation to their geographic position. Therefore, our data support the notion that studies on plant biodiversity should no longer be restricted to genetic aspects, especially in the case of vegetatively propagated plant species.