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Cultivated olive, a typical fruit crop species of the semi-arid regions, could successfully face the new scenarios driven by the climate change through the selection of tolerant varieties to salt and drought stresses. In the present work, multidisciplinary approaches, including physiological, epigenetic and genetic studies, have been applied to clarify the salt tolerance mechanisms in olive. Four varieties (Koroneiki, Royal de Cazorla, Arbequina and Picual) and a related form (O. europaea subsp. cuspidata) were grown in a hydroponic system under different salt concentrations from zero to 200 mM. In order to verify the plant response under salt stress, photosynthesis, gas exchange and relative water content were measured at different time points, whereas chlorophyll and leaf concentration of Na+, K+ and Ca2+ ions, were quantified at 43 and 60 days after treatment, when stress symptoms became prominent. Methylation sensitive amplification polymorphism (MSAP) technique was used to assess the effects of salt stress on plant DNA methylation. Several fragments resulted differentially methylated among genotypes, treatments and time points. Real time quantitative PCR (RT-qPCR) analysis revealed significant expression changes related to plant response to salinity. Four genes (OePIP1.1, OePetD, OePI4Kg4 and OeXyla) were identified, as well as multiple retrotransposon elements usually targeted by methylation under stress conditions.

Prokaryotes and unicellular eukaryotes such as yeasts or Plasmodium can synthesize folate de novo via the folic acid synthesis pathway (Figure 7.1). The steps of this pathway include:(i) the condensation of dihydropteridine pyrophosphate (DHPP) 1 with p-aminobenzoic acid (pABA) 2 by dihydropteroate synthase (DHPS, EC 2.5. 1.15) to form dihydropteroate (DHP) 3;(ii) the addition of glutamate to DHP by dihydrofolate synthase (DHFS, EC 6.3. 2.12) to form dihydrofolate (DHF) 4; and (iii) the reduction of DHF to form tetrahydrofolate (THF) 5, catalyzed by dihydrofolate reductase (DHFR, EC 1.5. 1.3)[1]. DHPS is thus an essential enzyme responsible for the de novo synthesis of the folate molecule. Tetrahydrofolate produced by this pathway is required for one carbon trans--fer reactions in the biosynthesis of a range of biomolecules, such as nucleotides and amino acids [2]. DHFR is the enzyme responsible for the maintenance of folate pools in their physiologic, reduced states. DHFR, in fact, is needed for the intracellular conver--sion of synthetic folic acid into the THF forms that can participate in folate/homocyst--eine metabolism. Reduction of DHFR enzymatic activity diminishes the THF pool inside the cells, affecting the level of folate coenzymes and thus purine and pyrimidine biosynthesis [3]. This may as well influence homocysteine levels and methylation pro--cesses, because methyl-THF is needed for the re-methylation of homocysteine to form methionine, thereby ensuring the provision of S-adenosylmethionine (SAM) necessary for most biological methylation processes [4]. In contrast to prokaryotes and

Olive (Olea europaea L.) is a perennial diploid species mainly clonally propagated and diffused in the Mediterranean area as one of the oldest tree crops (Loumou and Giourga, 2003; McKey et al., 2010; Zohary et al., 2012; Mousavi et al., 2017). As in many other allochthonous, hermaphrodite, wind-pollinated species, olive is characterized by a plentiful flowering, followed by a poor fruit set, which results in low yields (Cuevas and Polito, 2004; Ben Dhiab et al., 2017; Kassa et al., 2019). Environmental conditions, such as temperature, rainfall, and wind, may strongly affect flowering time, flowering intensity, and fertilization (Fernandez-Escobar et al., 2008; Haberman et al., 2017; Benlloch-González et al., 2018). Efficient pollination depends on many factors, such as the presence of exogenous compatible pollen, the duration of stigma receptivity, the number of pollen grains, pollen-ovule ratio, and stigma morphology (Cruden, 2000; García-Mozo et al., 2004; Pinillos and Cuevas, 2009; Rojo et al., 2016). Despite the importance of these factors, the main constraints responsible for the low fruit setting of olive are undoubtedly self-incompatibility (SI) and a high percentage of ovary abortion of some cultivars (Reale et al., 2009; Seifi et al., 2015). Self-incompatibility is one of the most effective systems adopted by flowering plants to prevent inbreeding and maintain a high diversity within species (Ferrer and Good, 2012). Within the sporophytic (SSI) and gametophytic (GSI) categories, several incompatibility systems have been reported, but only three of them have been characterized at molecular level, one for SSI (Brassicaceae) and two for GSI (one in Solanaceae, Plantaginaceae, and Rosaceae, and one for Papaveraceae) (Higashiyama, 2010). The recent discovery of the diallelic SSI system in olive and other related genera has provided evidence on the SI system operating in the Oleaceae family. In particular, it has been clarified that the inhibition of pollen tube growth takes place at the stigma, and it has been established that there are only two incompatibility groups, i.e., diallelic SI, such that olive cultivars are incompatible within groups and compatible between groups (Saumitou-Laprade et al., 2017a). In light of this evidence, it has become even more difficult to explain the self-fertility of some varieties, which has been confirmed by numerous studies (Seifi et al., 2012; Selak et al., 2014a; Breton et al., 2016; Marchese et al., 2016a). It is also now clear that pseudo-self-compatibility (PSC)--the failure to reject self-pollen despite the presence of a functional SI system--may occur in olive, and all previous reports on olive self-fertility need to be re-interpreted or ignored when not confirmed by paternity tests of seeds deriving from selfing. In this review, we hope to shed some light on the complexity of SI system in olive, particularly focusing on the evidence for PSC, providing a synthesis of the fragmented work on this topic and a new perspective on SI in olive, supported by experimental work from our laboratory

After uptake from soil into the root tissue, distribution and allocation of nitrate throughout the whole plant body, is a critical step of nitrogen use efficiency (NUE) and for modulation of plant growth in response to various environmental conditions. In legume plants nitrate distribution is also important for the regulation of the nodulation process that allows to fix atmospheric N (N2) through the symbiotic interaction with rhizobia (symbiotic nitrogen fixation, SNF).

Environmental conditions may strongly impact plant crop growth (Kachaou et al., 2010; Feller and Vaseva, 2014; Pandolfi et al., 2017). In particular, abiotic constraints, such as drought, soil salinity and extreme temperatures, which cause water depletion in cells, are responsible for a large proportion of losses in agricultural productivity (Bose et al., 2014). In order to overcome water shortages and to satisfy the increasing water demand for agricultural development, the use of water of low quality (brackish, reclaimed, drainage) that frequently has an high salinity level is becoming important in many countries (Chartzoulakis, 2005). In particular, plants under high salinity conditions are subject to significant physiological and biochemical changes, for example a marked decrease in photosynthesis rate and transport of salt ions from roots to shoots (Ben Ahmed et al., 2009; Anjum et al., 2011; Singh and Reddy, 2011; Goltsev et al., 2012; Abdallah et al., 2018). A major biochemical alteration, also induced by other types of stress, is the production of reactive oxygen species (ROS) (Gill and Tuteja, 2010; Boguszewska and Zagda?ska, 2012; Ozgur et al., 2013; Bose et al., 2014). An excess of ROS leads to lipid peroxidation, inhibition of enzymes, and modifications of nucleic acids (Proietti et al., 2013; Bose et al., 2014; Tedeschini et al., 2015). Under stress conditions, plants can nonetheless develop tolerance, that is, the ability to adequately survive, and often prosper, under an unfavorable environment, following a robust production of antioxidant enzymes (Ben Ahmed et al., 2009; Bhaduri and Fulekar, 2012; Keunen et al., 2013). Among these enzyme, superoxide dismutase (SOD), ascorbate peroxidase (APX), and glutathione reductase (GSH) are localized in chloroplasts and mitochondria (Pang and Wang, 2010; Del Buono et al., 2011; Proietti et al., 2013), whereas catalase (CAT) and guaiacol peroxidase (GPX) are generally present within microbodies and cytosol, respectively (Bhaduri and Fulekar, 2012; Hameed et al., 2013; Nath et al., 2016). Mechanistically, tolerance may also include osmotic adjustments at cellular level (Ayala-Astorga and Alcaraz-Meléndez, 2010). Some plants implement this process by increasing the amount of solutes and lowering the water potential of root cells, thereby counteracting the water outflow. These substances, reported as osmolytes, can accumulate in large amount, but do not generally interfere with enzymatic activities and cytoplasmic pH, due to their zwitterionic nature. Osmolytes commonly used by plants are sugars, alcohols, quaternary amines, betaine, glycine and proline (Warren, 2014). In this regard, the concentration of proline in leaves and roots was reported as a response by the olive tree to saline stress (Ayala-Astorga and Alcaraz-Meléndez, 2010; Hayat et al., 2012; Iqbal et al., 2014; Abdallah et al., 2018). In fact, proline facilitates water retention in the cytoplasm and, therefore, its concentration is indicative of response to saline stress (Ben Ahmed et al., 2009; Gupta and Huang, 2014). Cultivated olive (Olea europaea subsp. europaea var. europaea) is a long-living, evergreen, thermophilic species. In the Mediterranean basin where olive is mostly cultivated salinity is becoming a relevant problem due to high rates of evaporation and insufficient leaching (Mousavi et al., 2019). In addition in costal areas the need for water of good quality for urban use is increasing while there is a large amount of low quality water mostly saline (EC > 2.0 dS m-1) that can be use for irrigation (Chartzoulakis, 2005). Olive is considered as a moderately salt tolerant plant and the tolerance appear to be cultivar dependent (Rugini and Fedeli, 1990). The olive crop counts a very rich varietal heritage (Mousavi et al., 2017) but genotypic responses of olive to NaCl salinity have not been extensively investigated, and only few works have been published (Al-Absi et al., 2003; Chartzoulakis, 2005). In this context it's important to select cultivars that may give good performance when cultivated in soil with salinity problems or irrigated with saline water. Among the cultivars studied in the present work Arbequina and Koroneiki cultivars are the subject of increasing interest given their adaptability to super-intensive cultivation systems (Proietti et al., 2015). The identification of saline-resistant cultivars is of particular interest, especially for those cultivation systems, such as the super-intensive, which require large quantities of water as the availability of non-saline water will decrease dramatically in the future due to climate change. The purpose of this work was to study the behavior of different olive cultivars during saline stress by analyzing the activity of the GSH and CAT enzymes, the proline content and the plant growth parameters.

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Bacteria encode carbonic anhydrases (CAs, EC 4.2.1.1) belonging to three different genetic families, the alpha-, beta-, and gamma-classes. By equilibrating CO? and bicarbonate, these metalloenzymes interfere with pH regulation and other crucial physiological processes of these organisms. The detailed investigations of many such enzymes from pathogenic and non-pathogenic bacteria afford the opportunity to design both novel therapeutic agents, as well as biomimetic processes, for example, for CO? capture. Investigation of bacterial CA inhibitors and activators may be relevant for finding antibiotics with a new mechanism of action.

The field of thermophilic microbiology was born in the late 1970s with the pioneering work of Brock (Thermophiles biodiversity, ecology, and evolution. Springer, Boston, pp. 1–9, 2001) and dramatically expanded through the ’80s with the isolation of hyperthermophiles by Stetter (FEMS Microbiol Rev 18:149–158, 1996). The development of SSU rRNA phylogenetics revealed the complexity and diversity of prokaryotic phylotypes on biotopes widely differing in extreme conditions (e.g. spanning gradients of pH between 0 and 10 and temperatures from 60 °C to over 120 °C, respectively). Sites of volcanic activity all over the Earth’s surface and under the sea provide a variety of different environments for extremophilic microorganisms. Hot springs populated by hyperthermophiles (Topt > 65 °C), the majority of which belonging to the domain of Archaea, are very diverse and some of them show combinations of other extreme conditions, for example, acidic, alkaline, high pressure, and high concentrations of salts and heavy metals (Cowan et al. in Curr Opin Microbiol 25:97–102, 2015). Archaea inhabiting hot springs are considered to be the closest living descendants of the earliest living forms on Earth and their study provide insights into the origin and evolution of life (Woese et al. in Proc Natl Acad Sci USA 87:4576–4579, 1990; Olsen et al. in J Bacteriol 176:1–6, 1994). As with all studies of environmental microbiology, our understanding of the function of (hyper)thermophilic microbial consortia has lagged substantially behind. However, recent advances in ‘omics’ technologies, particularly within a system biology context, have made significant progresses into the prediction of in situ functionality (Cowan et al. in Curr Opin Microbiol 25:97–102, 2015). Most extremophilic microorganisms are recalcitrant to cultivation-based approaches (Amann et al. in Microbiol Rev 59:143–69, 1995; Lorenz et al. in Curr Opin Biotechnol 13:572–577, 2002); therefore, culture-independent metagenomic strategies are promising approaches to assess the phylogenetic composition and functional potential of microbial communities living in extreme environments (Lopez-Lopez et al. in Life 3:308–320, 2013). In addition, these approaches implement tremendously the access to enzymes from (hyper)thermophilic microorganisms that have important potential applications in several biotechnological processes. We report here on the state-of-the-art of the metagenomic surveys of different hot springs (T > 65 °C) (Table 5.1) and on the recent advance in the discovery of new hyperthermostable biocatalysts of biotechnological interest from metagenomic studies of these extreme environments.