Emerging evidence indicates that resolution of inflammation is a critical and dynamic endogenous process for host tissues defending against external invasive pathogens or internal tissue injury. It has long been known that autoimmune diseases and chronic inflammatory disorders are characterized by dysregulated immune responses, leading to excessive and uncontrol tissue inflammation. The dysregulation of epigenetic alterations including DNA methylation, posttranslational modifications to histone proteins, and noncoding RNA expression has been implicated in a host of inflammatory disorders and the immune system. The inflammatory response is considered as a critical trigger of epigenetic alterations that in turn intercede inflammatory actions. Thus, understanding the molecular mechanism that dictates the outcome of targeting epigenetic regulators for inflammatory disease is required for inflammation resolution. In this article, we elucidate the critical role of the nuclear factor‐κB signaling pathway, JAK/STAT signaling pathway, and the NLRP3 inflammasome in chronic inflammatory diseases. And we formulate the relationship between inflammation, coronavirus disease 2019, and human cancers. Additionally, we review the mechanism of epigenetic modifications involved in inflammation and innate immune cells. All that matters is that we propose and discuss the rejuvenation potential of interventions that target epigenetic regulators and regulatory mechanisms for chronic inflammation‐associated diseases to improve therapeutic outcomes. Innate immune system responds to stimulus  firstly; nevertheless, exaggerated immune response results in a variety of inflammation‐related diseases. Epigenetic modifications including chromatin remodeling, DNA modifications, RNA modifications, Histone modifications, noncoding RNA, and regulate multiple signaling pathways of chemokines and cytokines released by immune cells in inflammation response. Biotherapeutics targeting epigenetics regulators have been studied as an important clinically effective treatments for inflammation‐related diseases. This review mainly focuses on the inflammation‐related epigenetic mechanisms and summarizes the known contributions of targeting epigenetic modification to therapy in inflammation‐related diseases Figure .22.1Inflammation is the immediate response of the host tissues and cells to pathogens, harmful stimuli , or physical damage. The innate immune system responds rapidly to inflammation, which mainly comprises inducers, sensors, mediators, and effectors.During an inflammatory response, various inflammatory signaling pathways are activated and organized to regulate the expression of both pro‐ and anti‐inflammatory mediators. Based on previous studies, nuclear factor‐κB (NF‐κB) has been regard a prototypical proinflammatory signaling pathwayElevated production of IFNs during inflammatory response leads to upregulation of most typically canonical interferon‐stimulated genes (ISGs) expression. IFNs are composed of type I IFNs and the sole type II IFN (IFN‐γ) Figure , which hNod‐like receptor family pyrin domain containing 3 (NLRP3) is an intracellular receptor that senses foreign pathogens and endogenous danger signals, conducting the formation and activation of the NLRP3 inflammasome Figure , which iUnder normal circumstances, inflammation quickly ends after the clearance of infection and injurious agents. There is precise control of the complex networks of inflammatory pathways to limit tissue damage during inflammation, while continued activation of the immune system can lead to inflammatory dysregulation. Growing evidence suggests a close link between inflammation and many chronic health conditions, including autoimmune diseases, ND, viral infections such as coronavirus disease 2019 (COVID‐19), and cancer.2.2Chronic inflammatory diseases make up a burden of human health. Although the origins of some chronic diseases remain to be clarified, the dysregulated inflammatory response has been evidenced to lead to pathological conditions.2.2.1SLE is a condition in which the immune system attacks healthy cells and tissues throughout the body. The pathogenesis of SLE is characterized by exaggerated immune responses and loss of tolerance against self‐antigens. Antibody production and defective elimination, complement and cytokine activation, and tissue deposition of immune complex lead to clinical manifestations ranging from mild fatigue and joint pain to severe, life‐threatening organ damage.The dysregulation of apoptosis and nuclear debris clearance contribute to an increase in self‐antigen exposure. Accumulated apoptotic debris can trigger TLRs and nucleic acid sensors that facilitate their clearance by cells in the reticuloendothelial compartment. TLRs, such as TLR3, TLR7, TLR8, and TLR9, are transferred into the endoplasmic reticulum by the trafficking protein unc‑93 homologue B1 (UNC93B1). TLR9 is a receptor for DNA containing unmethylated CpG sequence motifs. In plasmacytoid dendritic cells (DCs), TLR9 and IRF7 induce a strong IFN immune response in early endosomes, whereas TLR9 and NF‐κB drive a proinflammatory cytokine response in late endosomes.Accumulating evidence suggested that the NLRP3 inflammasome was hyper‐activated in SLE. In macrophages and peripheral blood mononuclear cells (PBMCs) of patients with SLE, dsDNA and anti‐dsDNA autoantibodies have been reported to upregulate NLRP3 and caspase‐1 expression, improving the production of IL‐1β.2.2.2RA is one of the most prevalent chronic inflammatory diseases that endanger human health, its predisposing factors mainly include positive family inheritance and environment factors, such as smoking and low socioeconomic status or educational attainment.Early in the disease course of the RA, bone loss occurs due to continued direct exposure of bone and cartilage to the inflammatory microenvironment. This progress is driven by the induction of bone‐resorbing osteoclasts by autoantibodies, leading to the first structural changes in the pre‐disease stage and exacerbated by the action of proinflammatory cytokines.2.2.3Neuroinflammation initially protects the brain by removing or inhibiting diverse pathogens, promoting tissue repair and removing cellular debris. However, sustained inflammatory responses have a detrimental effect on health.The CNS consists of two categories of cells: neurons and glial cells.Amyloid‐β (Aβ) deposition in the brain is one of the starting events in AD. Aβ activates the NF‐κB pathway in astrocytes, leading to increased release of complement C3, which successively acts on C3a receptors on neurons and MG, resulting in neuronal dysfunction and microglial activation.2.3The COVID‐19 pandemic, caused by SARS‐CoV‐2, has spread worldwide.For RNA viruses such as SARS‐CoV‐2, the innate immune response is initiated through the engagement of PRRs by viral single‐stranded RNA (ssRNA) and double‐stranded RNA (dsRNA). Upon PRR activation, downstream signaling cascades trigger the secretion of cytokines.Similarly to SARS‐CoV, spike (S) protein is expressed on the surface of SARS‐CoV‐2 virus particles, and several receptors have been showed for the interaction of spike (S) protein with host cells, including angiotensin‐converting enzyme (ACE2). ACE2 has a crucial role because loss of pulmonary ACE2 function is associated with acute lung injury (ALI), and virus‐induced ACE2 downregulation may be important for disease pathology.2.4Inflammation predisposes patients to the development of cancer and facilitates all stages of tumorigenesis. Approximately 15%–20% of all cancer cases develop infection, chronic inflammation, or autoimmunity at the same tissue or organ site. Inflammation has a great impact on the composition of the tumor microenvironment (TME) and particularly on the plasticity of tumor cells, as well as surrounding stromal and inflammatory cells, engages in orchestrated interactions to shape the inflammatory TME.Several pathways are broadly implicated in inflammatory processes and are closely linked to tumorigenesis in multiple tissues. Part of the complexity and differences in NF‐κB function between different tumor types is because of its crosstalk with other transcription factors and signaling proteins. NF‐κB promotes STAT3‑mediated expression of genes encoding different inflammatory mediators in immune cells of the TME.While cellular senescence intrinsically suppresses the tumorigenesis of preneoplastic cells, senescent cells show widespread changes in gene expression and chromatin organization. These changes include the secretion of numerous proinflammatory cytokines, chemokines, growth factors, and proteases, which is a feature known as the senescence‐associated secretory phenotype (SASP). The SASP produced by senescent cells can actually extrinsically promote tumor growth, relapse, and metastasis.33.1Chromatin structure dynamics is necessary for regulating gene expression, and is mostly controlled by a group of ATP‐dependent chromatin remodeling regulators which have an ATPase domain of catalytic subunits. ATP‐dependent chromatin remodeling regulators use energy from ATP hydrolysis to modulate DNA translocation on nucleosome and histone composition, regulate the assembly, disassembly, and rearrangement of nucleosomes on chromatin.yeastDifferent chromatin remodeling factors have similarities in protein structure and enzyme activity, and each has its own specificity. Based on functional analysis, ATP‐dependent chromatin remodeling factors can be roughly divided into four subfamilies: CHD (chromo domain helicase DNA binding), ISWI (imitation switch), SWI/SNF (switch/sucrose non‐fermentable), and INO80.3.2DNA methylation has precision regulatory functions in various biological processes, which can be stably inherited through meiosis, persists in mitosis, and is extremely important in maintaining genome integrity.5‐Methylcytosine (5mC) catalyzed by DNMT Figure  was the As an important epigenetic mechanism, DNA methylation is implicated in various biological processes of immune cells, including cell activation, proliferation, differentiation, and apoptosis, all of which are associated with the pathogenesis of inflammatory diseases.As reviewed by Schmidl et al.,Demethylation in mammals is predominantly mediated by the TET methylcytosine dioxygenases family—namely TET1, TET2, and TET3.Additionally, N6‐methyladenine (6mA), as a modification of distribution universally in the human genome, has drawn widespread attention since 6mA in various eukaryotic genomes was confirmed in 2015.3.3The fundamental unit of chromatin is nucleosome, and the core region of nucleosome comprises histone octamers  that are enriched by abundant PTMs. Histone PTMs not only occur in the terminal tails, but on the lateral surface of nucleosome core regions.3.3.1Due to its existence in three distinct states on both arginine  and lysine  residues, methylation on histone tails represents a complex and more subtle chromatin modification than acetylation.Lysine can be mono‐, di‐, or trimethylated by six major classes of histone lysine methyltransferase (KMT1‐6). KMT1 family that is in chargeof methylation of histone H3 at lysine 9 (H3K9) contains four members, including G9a, GLP,TNF‐α, IL‐6, NOS2, MHC‐II, and a regulator of MHC‐II expression (CIITA). RelB a subunit of NF‐κB, directly binds and recruits G9a to IL‐1β promoter in THP‐1 endotoxin tolerant macrophages, forming a complex with HP‐1 and promoting gene silencing through H3K9 methylation.TNF‐α, IL‐1α, IL‐1β, CCL‐4, and CXCL10.Histone methylation contributes to binding effector molecules with DNA in response to various cellular signals, which exert vital roles in gene transcription, repair, and replication processes.Foster et al.Recently, a plethora of research studies have drawn attention for the potential role of histone demethylases in catalyzing the removal of methyl groups from lysine and arginine residues in the regulation of inflammatory gene expression.3.3.2Histone acetylation has a powerful function in regulating chromatin structure through neutralizing positively charged lysines and recruiting bromodomain‐containing proteins (BRDs) to promote the opening of chromatin, then allowing access to transcription factors and other transcription coactivators.HDACs have been implicated in the regulation of proinflammatory genes, and they reverse the activity of HATs by removing acetyl groups from histones, resulting in the condensation of chromatin and the repression of inflammatory gene transcription Figure . For ins3.3.3Histone ubiquitylation, like other modifications, also plays an important role in regulating gene transcription and chromatin structure in mammals.Histone ubiquitination is also a reversible epigenetic modification, deubiquitinating enzymes are grouped into six subfamily members in human genomes: ubiquitin C‐terminal hydrolases, MCP 1‐induced protein, Josephins, JAB1/MPN/MOV34 family (JAMMs), ubiquitin‐specific proteases (USPs), and ovarian tumor proteases.Mono‐ubiquitination exerts a critical role in transcriptional regulation, protein translocation, and DNA damage signaling. Several studies have revealed that mono‐ubiquitination of histone 2A (H2Aub) and histone 2B (H2Bub) were widely involved in the development of inflammation progression. Whereas, H2Aub is typically correlated with gene silencing, and the latter is more often associated with transcription activation.The mono‐ubiquitination of histone H2B at lysine 120 (H2B K120ub1) mediated by the ubiquitin ligases RNF20 and RNF40 was shown to play context‐dependent roles in the development of inflammation. Koskinsky et al.3.3.4trans, and promotes acetylation on histone H3 at lysine 27 (H3K27ac) at mESC enhancers, suggesting that H3.3 plays an important role in guiding gene transcription.Histone phosphorylation take place on serine, threonine, and tyrosine residues,3.4All known RNA species can be the targets of modifications, with ribosomal RNA (rRNA) and transfer RNA (tRNA) being the most modified.Recently, it was found that the levels of m6A and METTL3 expression were upregulated in human dental pulp cells (HDPCs) upon LPS stimulation. Similarly, deletion of METTL3 in HDPCs following LPS treatment increases the expression of MyD88S, leading to a decrease in the expression of inflammatory cytokines and suppressing the activation of the NF‐κB and MAPK signaling pathways.Similar to modifications that occur on DNA, RNA can be methylated at position 5 of cytidine residues. RNA 5mC has been identified and found on mRNA, rRNA, ncRNA, tRNA, and enhancer RNA (eRNA). The enzymes are responsible for 5mC modification of RNAs including seven members of the NOL1/NOP2/SUN domain (NSUN) family, namely NSUN1 to NSUN7, and DNMT‐like 2 (DNMT2). 5mC in RNA can be oxidized by Tet‐family enzymes to 5hmC.3.5Based on the Encyclopedia of DNA Elements (ENCODE) project, ∼80% of the human genome is transcribed into various ncRNAs,3.5.1Caenorhabditis elegans in 1993 and the first mammal miRNA (let‐7) was discovered in 2000.miRNAs, a novel class of endogenous ncRNAs of 18–25 nucleotides, are involved in the regulation of messenger RNA stability and translation at the posttranscriptional level in eukaryotic organisms.Baltimore's group first reported that PAMP recognition, such as microbial cell wall components or viral nucleic acids or bacteria, was coupled with the expression changes of various miRNAs.Escherichia coli increased the expression of miR‐146a/b, miR‐155, and miR‐132.Upon stimulation of cell surface receptors, such as TLRs, tumor necrosis factor receptors, IL‐1 receptors and receptor for advanced glycation endproducts, miR‐146a and miR‐146b, which are located on chromosomes 5 and 10 in human, respectively, are both upregulated.miR‐155, similar to the function of miR‐146 in TLR‐induced inflammation, was also considered a feedback negative regulator of the TLR‐NF‐κB axis signaling pathway in human and murine macrophages and monocytes Figure 218. MorIn a follow‐up study, further miRNAs were shown to have well‐documented functions in TLR‐induced inflammatory responses Figure . BesidesmiR‐203, the first skin‐specific miRNA, is involved in regulating various physiological and pathological processes Figure . It has 3.5.2cis virus and those that leave the site of transcription and carry out their functions throughout the cell in trans. Interestingly, the interactions of lncRNAs with mRNAs can significantly affect mRNA stability, translation or isolation. Additionally, the interaction between lncRNAs and multiple proteins can strengthen the assembly of protein complex or weaken the interaction between proteins. Recent studies have even revealed that lncRNAs could act as precursors of miRNAs or circRNAs.LncRNAs, which are broadly distributed in mammals, are the largest portion of the noncoding transcriptome with little or no protein translation capacity. LncRNAs are over 200 nucleotides and have been proposed to participate in a variety of biological processes through complicated mechanisms.cis and in trans by establishing agonistic and antagonistic interactions with proteins.Similar with miRNAs, accumulating evidence has indicated that lncRNAs were involved in both inflammatory pathways and immune reactions. Unlike the intensively studied miRNAs, most lncRNAs seem to control various cellular processes ranging from the cytosolic receptor signaling cascade to the opening or closing of local chromatin structures in Within the IL‐1β locus, for instance, the lncRNAs IL‐1β‐eRNA and IL‐1β‐RBT46 ‐dependent pathway Figure , which mInflammatory stimulation, for example, with diverse reagents, such as LPS and IFN‐γ, results in the induction of lincRNA‐p21 via the p53‐dependent pathway in microglial cells.4With the continuous development of epigenetics, research into epigenetic‐related drugs has made substantial progress. At present, numerous small‐molecule inhibitors or activators involving epigenetic regulation pathways are being developed for clinical therapy. The target of epigenetic drugs mainly includes DNMTs, DNA demethylases, HDACs, HATs, histone methyltransferases (HMTs), histone lysine demethylases (KDMs), and others.4.1DNMTs and DNA demethylases coordinate regulation of the genome's methylation status.With the deepening of clinical research, these nucleotide analogs exhibit many threats, including unstability chemically and metabolically, lack of specificity, and significant toxic side effects. Based on this phenomenon, more stable and less‐toxic cytidine analogs and their prodrugs: second‐generation DNMTis are designed and used for the treatment of inflammatory diseases. The second‐generation DNMTis comprises zebularineAdditionally, analogs of non‐nucleosides have been discovered over the last few decades to resolve the poor bioavailability, chemical instability under physiological conditions, and lack of selectivity of nucleoside analogs. SGI‐1027 is a quinoline‐based compound with antioxidant activity against DNMT1 Figure , DNMT3A,4.2Therapeutic targeting of chromatin modifiers involved in the regulation of specific modulators of innate immunity and their specific inhibitors and activators will be a potential therapeutic strategy for inflammation‐related diseases. Research into the multifaceted effects of histone modification inhibitors has long positioned these drugs as promising drug candidates for the treatment of cancer, autoimmunity, neurodegenerative, and infectious diseases.4.2.1Based on extensive research targeting epigenetic regulators, HDAC inhibitors have aroused wide attention. HDAC inhibitors enhance the level of acetylation modification of histone or nonhistone lysine residues by inhibiting the activity of HDACs, controlling the tightness of DNA wrapping around histones, and making DNA wrap around histones more tightly, which leads to this DNA not being easily accessible to gene transcription factors Figure . HDAC inThe FDA has also approved some HDAC inhibitors for clinical therapy. The first HDAC inhibitors approved for use in treating cutaneous T cell lymphoma (CTCL) were vorinostat (SAHA) and romidepsin4.2.2+ and IFN‐γ+CD8+ T cells involved in antitumor immunity.+ and CD8+ T‐cell H3K79me2 levels. Based on the findings of Kealy et al.,Studies have shown that histone methylations have undergone dramatic changes in inflammation‐related diseases including tumors,+ T cells.With the continuous development of epigenetic mechanisms and experimental techniques, various inhibitors of histone methyltransferases and histone demethylases have been discovered.4.3The m6A RNA modifications are the most common RNA modifications in eukaryotes, and involved in multiple aspects of RNA metabolism such as splicing, stability, structure, translation, and export.METTL3 and METTL14 inhibits the growth of cancer cells. For example, Cui et al.Mettl3‐ or Mettl4‐knockout glioblastoma stem cells (GSCs) had larger tumors compared to control group. After Mettl3 overexpression or Fto inhibition, GSC growth and self‐renewal are promoted and mice survived longer.Mettl3 and Fto.METTL3 and METTL14, m6A writers, play an important functions in tumors and the immune system and serve as potential therapeutic targets. Overexpression of FTO increased m6A methylation in PD‐1, CXCR4, and SOX10, causing increased RNA decay through the m6A reader YTHDF2. Consequently, FTO depletion enhanced the response of mouse melanoma cells to IFN‐γ and anti‐PD‐1 therapies.Alkbh5‐deficient mice, but mature neurons were reduced,Alkbh5 deficiency affects the proliferation and differentiation of neuronal progenitors.FTO and ALKBH5, two “erasers” of m6A, also play an important role in regulating inflammatory diseases. As reviewed by Garbo et al.,Ythdf1‐deficient mice, with stronger activation of antitumor CD8+ T cell.+ T‐cell responses against tumors were observed in Ythdf1‐deficient mice. The efficacy of PD‐L1 checkpoint blockade was also enhanced.m6A “readers” which recognize m6A modifications, are also important for the regulation of inflammatory diseases. The initiation of T‐cell immune responses is enhanced in 4.4Regulatory ncRNAs, including small noncoding RNAs, lncRNAs, and circRNAs, have gradually been discovered in the past few years for their mechanisms of action in inflammatory diseases. As therapeutic targets and promising biomarkers for inflammatory diseases, they have aroused extensive attention. RNA‐based drugs are designed and used in treating inflammation‐related diseases. miRNAs negatively regulate mRNA expression, thereby interfering with posttranscriptional regulation in various diseases. Likewise, lncRNAs are involved in cellular processes by affecting epigenetic, posttranscriptional, and translational pathways.Currently, 40 varieties of SARS‐CoV‐2 miRNAs and their regulatory targets have been identified.5Chronic inflammation causes various inflammation‐related diseases during the immune response of cells and the body. Epigenetic modifications have been accordingly changed with the occurrence and development of inflammation reaction. In turn, epigenetic alterations intercede inflammatory actions. Therefore, it is expected to reverse chronic inflammatory environment in the body via targeting these epigenetic regulators. Although clinical epigenetic drugs are still in its nascent stage, several epigenetic drugs have been approved, and a series of epigenetic marks exist or are under development for disease diagnosis, treatment response, and prognosis. Tumor therapy is currently the focus of clinical epigenetics, but numerous clinical trials and preclinical studies are also being conducted in other areas of medicine beyond cancer . Together, targeting epigenetic regulators would be an effective strategies for inflammatory diseases treatment by using predictive biomarkers or novel screening methods. Compared with traditional chemotherapeutics, epigenetic drugs show great advantages in the diversity and feasibility of biotherapeutics, but their clinical application hardly utilized. There are still many problems that are difficult to solve. We summarize as follows.First, based on the complex regulation of epigenetic modification, this therapeutic approach may be limited by its lack of specificity. Currently, FDA‐approved drugs, located at multiple genomic loci, play an equal role in re‐expression of silenced genes, which potentially can reactivating nonsense sequences in the genome, but have no differential effect on their epigenetic marks. Second, interactions between epigenetic factors give rise to the “promiscuous” nature of specific epigenome‐modifying enzymes. Multiple PTMs of histones can target the same amino acid residue, resulting in competitive antagonism at these PTMs sites. Therefore, targeting inhibition or activation of one PTM may lead to changes in the other modifications, thus affecting other cell functions. As an example, targeting HDAC inhibitors, such as vorinostat and sodium butyrate, can also alter histone methylation and the binding of chromatin remodeling factors. In addition, HDAC inhibitors can also alter CpG methylation at specific promoters by affecting DNMT activity. One reason for this interaction is that epigenetic modifiers form complex with other chromatin‐associated proteins. As a result, interventions in one partner may have unforeseen consequences for other partners. Epigenetic drugs frequently suffer from these drawbacks, which are often accompanied by significant side effects. Third, the majority of epigenetic drugs have been designed to target DNA methylation, histone methylation, and histone acetylation. Although the role of some epigenetic modifications in inflammation‐related diseases has been elucidated, candidate drugs targeting in chromatin openness, RNA modification, and DNA modification remain unidentified. Therefore, targets for these modifications will be an underlying clinical strategy for inflammatory disease. Fourth, the use of epigenetic drugs may benefit some patients with inflammatory diseases, but liking cancer chemotherapy, not all patients can benefit from monotherapy. So far, the FDA and other regulatory agencies have rarely approved epigenetic drugs for advanced cancer and inflammation‐related diseases due to insufficient efficacy, except for a few types of solid tumors. Therefore, it is possible to improve the therapeutic antitumor efficacy of epigenetic drugs by combining with other drugs. As the field of immunology develops, it is believed that future consortium of therapeutic modalities such as epigenetic therapy and immunotherapy are rapidly emerging as new paradigms for the treatment of inflammation‐related diseases.The authors declare they have no conflicts of interest.S.Z., Y.M., L.Z., and J.H. conceived, edited, and wrote this manuscript. L.Q., H.W., D.S., B.Z., K.‐M.C., and J.H. revised this manuscript. All authors have read and approved the final manuscript.The author declare that ethics approval was not needed for this study.


Olea europaea L.) orchards are fertilized, mostly with the macronutrients nitrogen (N), phosphorus (P) and potassium (K). The effects of different application levels of these nutrients on olive oil composition and quality were studied over 6 years in a commercial intensively cultivated ‘Barnea’ olive orchard in Israel.Intensive olive (Oil quality and composition were affected by N, but not P or K availability. Elevated N levels increased free fatty acid content and reduced polyphenol level in the oil. Peroxide value was not affected by N, P or K levels. The relative concentrations of palmitoleic, linoleic and linolenic fatty acids increased with increasing levels of N application, whereas that of oleic acid, monounsaturated‐to‐polyunsaturated fatty acid ratio and oleic‐to‐linoleic ratio decreased.Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.These results indicate that intensive olive orchard fertilization should be carried out carefully, especially where N application is concerned, to avoid a decrease in oil quality due to over‐fertilization. Informed application of macronutrients requires leaf and fruit analyses to establish good agricultural practices, especially in view of the expansion of olive cultivation to new agricultural regions and soils. © 2022 The Authors.  Olea europaea L.) oil is a major component of the Mediterranean dietOver the last few decades, the oil olive industry has been shifting from non‐irrigated, extensive olive orchard management to irrigated, intensive practices.et al.et al.One of the main health‐related attributes of olive oil is its high levels of monounsaturated fatty acids (MUFA) – especially oleic acid – which comprises 55–85% of its fatty acid content. The health‐promoting properties of oleic acid have been widely studied, and its benefits have been established. For instance, a recent systematic review and meta‐analysis suggested that oleic acid supplementation exerted a significant reduction in the concentration of C‐reactive proteins in the blood.−1. PP also affect the sensory properties of olive oil, determining its bitterness and pungency.Free fatty acid (FFA) content is a primary parameter for olive oil quality, determining its market value and its suitability for human consumption. Extra virgin olive oil (EVOO) is classified as such when its FFA content is less than 0.8%, whereas non‐edible olive oil contains above 3.3% FFA. The PP content of virgin olive oil (VOO) is another important quality parameter. These are strong antioxidants that have multiple roles in olive oil; in addition to protecting PUFA from oxidation, thereby extending the oil's shelf life, PP from the consumed oil serve as antioxidants in human tissues.et al.et al.et al.The increasing demand for high‐quality olive oil is an important factor in assessing the overall performance of olive orchards. Olive oil quality, especially FFA levels and PP content, but also fatty acid composition, is closely related to agricultural practices such as irrigation,−1. From 2007 until 2010, the orchard was alternately irrigated with brackish water and recycled wastewater according to water availability. From 2010, the orchard was irrigated with fresh water . Trees were generally trained to form a cup shape suitable for mechanical trunk shaking, with a single trunk of about 70 cm in height and three to five leaders per tree. The orchard was monitored and treated regularly for pests, especially the olive fly (Bactrocera oleae). Experimental plots consisting of at least 12 trees  were allocated over 3 ha of the orchard. Two uniform trees in the center of each plot were selected for sampling and measurements. The experimental setup was randomized blocks with seven plots per treatment, giving 14 trees (replicates) per treatment. Trees were irrigated twice a week through a drip system (one drip line per row of trees with 1.3 L h−1 drippers spaced at 0.75 cm). Seasonal irrigation was initiated after three consecutive weeks without precipitation  and stopped after the first significant rainfall . Irrigation quantity was according to local commercial practice, determined according to the return of potential evapotranspiration, calculated by a modification of the Penman–Monteith method,The experiment was carried out over 6 years (2011–2016) in a commercial intensively cultivated olive orchard located on the southern coastal plain of Israel at an elevation of 80 m above sea level, near Kibbutz Negba . The average annual temperature and rainfall at the site were 20.1 °C and 490 mm, respectively. Orchard soil texture varied between loam and clay loam. The orchard was planted in 2007 with cv. Barnea trees,75) received the commonly applied P and K levels ; the P0 treatment received no P, and 150 and 250 kg ha−1 y−1 of N and K, respectively, etc. The common commercial practice in the region is to apply 150, 35 and 250 kg ha−1 y−1 of N, P and K, respectively. In all treatments, the ratio of N forms was NH4+:NO3− = 1:1.The experiment included four N, two P and two K fertilization levels. The amounts of each nutrient applied per treatment are specified in Table −1 y−1  treatment appeared to be in poor condition, exhibiting severe visual N‐deficiency symptoms. Therefore, in order not to lose the trees completely, in the 2015 and 2016 seasons, their N fertilization was adjusted to 40 kg ha−1 and the treatment was redesignated as N0,40. Each fertilization level (treatment) had a separate computer‐controlled and monitored irrigation and fertilization application system, including a tailored liquid fertilizer source container. The liquid fertilizers were prepared according to each treatment's specifications and supplied by Israel Chemicals Ltd . Each fertilizer was compiled such that annual application per hectare included, in addition to the specified N, P and K levels, 3.8 kg iron (Fe), 1.9 kg zinc (Zn), 940 g manganese (Mn), 140 g copper (Cu) and 100 g molybdenum (Mo). To ensure accuracy, irrigation solution was sampled from the drippers every 2 weeks, from two plots for each treatment, and nutrient concentrations were confirmed. In the season prior to the initiation of the experiment (2010), the orchard was fertigated constantly during the irrigation season with annual amounts of 250 kg N ha−1, 175 kg K ha−1 and no P.Differential fertilization treatments were initiated in June 2011 and were implemented over a period of six consecutive years, up to and including the 2016 season. During the experiment, trees in the plots receiving the 0 kg N haet al.Fruit was harvested upon ripening (‘50% black’), between October and November, from individual trees by a mechanical trunk‐shaker with simultaneous rod beating. Fruit yield per tree was weighed and a 3 kg subsample was taken to the laboratory and kept in a cool environment, in aerated boxes. Oil was cold‐extracted from the fruit within 24 h of harvest using a laboratory mill , according to the procedure described by Ben‐David −1 oil), was determined with a UV–visible spectrophotometer  at a wavelength of 735 nm, using Folin–Ciocalteu reagent. Fatty acid composition was determined in the years 2014–2016 by gas chromatograph model 6890 N with flame ionization detection and a DB‐23 capillary column . Helium, nitrogen and air were employed as carrier gases at a flow rate of 1 mL min−1 and the split ratio was 1:50. The injection volume was 1 μL. The initial oven temperature was 165 °C, maintained for 15 min at a heating rate of 5 °C min−1 to 200 °C, and then elevated to 230 °C at 25 °C min−1 and held at this temperature for 12 min. The temperature of the injector and flame ionization detector was set to 240 °C. The total running time was 35.2 min. The fatty acid peaks were identified using Agilent Technologies software 5988‐5871EN and results were identified by comparing retention times with standard compounds. The relative content of the fatty acid was recorded as peak area percentage. The relative concentrations of eight of the major fatty acids in olive oil were determined: palmitic (C16:0), palmitoleic (C16:1), stearic (C18:0), oleic (C18:1), linoleic (C18:2), linolenic (C18:3), arachidic (C20:0) and eicosenoic (C20:1), which made up over 94% of the total lipids. In addition, the ratios of unsaturated fatty acids (UFA) to saturated fatty acids (SFA) and those of MUFA to PUFA were calculated.FFA levels and PV were determined using International Organization for Standardization (ISO) analytical methods 660 and 3960, respectively. FFA level was expressed as percent oleic acid, and PV was expressed as millimoles of active oxygen per kilogram of oil. Phenolic compounds were isolated from a solution of oil in hexane by double‐extraction with methanol–water . PP content, expressed as tyrosol equivalents , digested with a mix of sulfuric acid and hydrogen peroxide, and the supernatant was analyzed. N and P contents were determined by means of an auto‐analyzer , and K content was determined with a Perkin‐Elmer AAnalyst 200 atomic absorption spectrophotometer (Milwaukee WI).A paste sample was taken after crushing the olives and before oil extraction. The sample was dried at 70 °C until its weight stabilized, milled and analyzed for N, P and K contents, using the same procedure described for leaves. Results were presented as percentage of dry matter in both cases.P ≤ 0.05.Data were analyzed by two‐way analysis of variance (ANOVA) of year and treatment, using JMP software version 14 . Significant differences were determined by Tukey–Kramer honest significant difference test at As found previously,Table Two‐way ANOVA revealed no interaction between year and treatment and, therefore, only the treatment effect is presented.0,40) to 1.14% (VOO) with application of 300 kg ha−1 y−1. PV decreased slightly and consistently with increasing N fertilization , 274 mg kg oil−1, whereas the lowest PP level, about 163 mg kg oil−1, was obtained with the highest N application level.The highest PP content Fig.  in the oThe relative concentrations of the eight fatty acids measured in this work are presented in Fig. 0versus P80 treatment.FFA content, PV and PP content of the P and K fertilization treatments for the years 2011–2016 are presented in Table The relative concentrations of the fatty acids measured in this work for the P and K treatments are presented in Table et al.As intensive cultivation of oil olives expands, a deeper understanding of the effects of irrigation and fertilization on oil yields and quality is emerging.0,40 and N75, leaf N concentrations reached deficiency levels after 5 years 300 with a leaf N concentration of 17.93 g kg−1. The P0 treatment was on the border of deficiency after 5 years (0.98 g kg−1) and the K0 treatment did not reach deficiency level. However, P and K leaf concentrations in the P and K fertilized treatments were significantly higher.The strong positive relations between N, P and K application levels and their concentrations in leaf and fruit dry matter Table  enabled et al.,−1) was accompanied by yield reduction. Similar results were obtained in a pot experiment carried out using the same olive cultivar.−1 y−1. Higher N application rates did not increase oil yieldet al.et al.,et al.In Haberman et al.et al.,et al.et al.et al.,SFA  concentrations were not affected by N application levels. This is in agreement with the results of Dag et al.et al.The effect of P application on olive tree performance has been demonstrated only recently.et al.,et al.et al.The relative concentrations of fatty acids in the oil in our experiment were not affected by P fertilization levels, in contrast to Dag et al.0 treatment, with no supplemental K fertilization for 6 years, leaf K concentrations did not drop below the accepted threshold value for deficiency of 0.8%. This, again, is probably due to the large K reserves in the soil, which become available as K depletion proceeds.et al.,Of the three macronutrients studied in the current work, K was the only one to show higher concentrations in olive fruit paste compared to leaves Table . The effOlive oil quality is an important parameter market‐wise as well as health‐wise. We found that in intensive olive orchards N fertilization levels play an extremely important role in the resultant oil quality, whereas P and K are less important. Controlled N fertilization of olive orchards and of many other crops has recently been reviewed, highlighting the vast effects on the environment and food security.Agricultural soils contain varying amounts of macro elements, and it is therefore difficult to derive general recommendations for fertilization to optimize oil yield and quality. Leaf analysis and, in the future, maybe also fruit analysis should serve as a useful diagnostic tool for tree nutritional status and the required corrective fertilization steps, especially as olive cultivation expands to new agricultural regions and soils.IZ: conceptualization, methodology, formal analysis, investigation, writing. UY: conceptualization, methodology, investigation, funding acquisition, writing – review and editing. AD: conceptualization, investigation, writing – review and editing. RE: conceptualization, investigation, writing – review and editing. ABG: conceptualization, investigation writing – review and editing. LQ: investigation, chemical analysis, data analysis, writing. ZK: conceptualization, investigation, writing – review and editing.The authors declare no conflicts of interest.


Paroxysmal dysarthria and ataxia (PDA) is a rare neurological manifestation of stereotyped attacks of sudden ataxic symptoms lasts for few seconds to minutes. We report a case of PDA in a 61‐year‐old male with a solitary homogenously enhancing solitary midbrain lesion and positive HLA‐B51 (Allele 2), controlled with lacosamide. Due to paucity of isolated midbrain lesion reports in neuro‐Bechet's disease presenting with PAD, neurologists and physicians must have a low threshold to explore HLA‐B51 as part of the diagnostic workup for isolated midbrain lesion, despite the absence of other NBD diagnostic criteria. No specific aggravating factors and episodes did not exhibit diurnal variability. No history of preceding illness, trauma, and no constitutional symptoms were present. A review of systems revealed no extra‐neurological manifestations, no mouth, genital ulcers, or blurred vision and no relevant family history, namely PDA, cerebellar ataxia, stammering, or stuttering.Initial vitals were within normal parameters, temperature of 36°C, respiratory rate of 19 breaths per minute, blood pressure of 120/62 mmHg, oxygen saturation of 98%, and weight of 75 kg. His neurological examination was significant only for right dysmetria, dysdiadochokinesia, and inability to tandem. During the consultation, episodes were witnessed of 10–20 s stereotyped ataxic dysarthria with dysprosody and none unintelligible, pointing toward paroxysmal ataxia and dysarthria (PAD). Neuropsychological assessment was unrevealing, with normal Mini‐Mental State Examination (MMSE) 29/30.Initial workup; MRI head with contrast showed 5 × 8 × 6 mm homogenously enhancing midbrain lesion, a midline lesion with suspicious thickening and increased enhancement of left third cranial nerve that raised the suspicion of underlying neoplastic vs. inflammatory etiology, such as lymphoma, glial tumors, or neuro‐sarcoidosis or neuro‐Behcet's disease Figure . SubsequLumbar puncture was performed and showed no pleocytosis, normal cerebrospinal fluid (CSF) biochemistry, culture for bacteria, and nPCR assays for a broad panel of viruses were negative. No malignant cells were visible on cytology, and flow cytometry was not done due to paucity of cells, and no oligoclonal bands were detected. Blood tests were within normal parameters, including autoimmune workup and angiotensin‐converting enzyme level (ACE) 61 U/L .Despite the possible differential of glial tumor or lymphoma, a brain biopsy was deferred and not advised by the neurosurgery team due to high risk. Due to anatomical location and radiological features, a possible diagnosis of neuro‐Behcet's was considered. The ophthalmology examination was normal, with no evidence of anterior or posterior uveitis on slit‐lamp examination. Further HLA‐B51 (Allele 2) analysis was requested .The patient was started on pulsed steroids with one gram of intravenous methylprednisolone for 5 days with a tapered dose (IVMP), which did not note any significant improvement in the frequency of his paroxysmal ataxia and dysarthria episodes. He was started on 200 mg carbamazepine and discharged home with regular follow‐ups in the General Neurology clinic.On 4‐week follow‐up, the patient developed fever, deranged liver function test (LFT), and severe skin maculopapular rash on more than one‐third surface area, suggestive of anticonvulsant hypersensitivity syndrome (AHS), significant enough to discontinue carbamazepine and was switched to baclofen. His PDA episodes were not adequately under control; hence, a trial of lacosamide 50 mg BID controlled his PDA episodes.On 5‐week follow‐up, HLA‐B51 results reported positive, raising the diagnosis of PAD in neuro‐Behcet's with atypical radiological and clinical findings. Per Behcet's syndrome Japanese Ministry of health and welfare criteria, our patient has one minor criterion due to a symptomatic central nervous system (CNS) lesion. A 3‐month follow‐up MRI head showed slight interval regression in the midline midbrain lesion size to 4 × 7 × 5 mm in maximum AP, with decreased perilesional edema Figure .3Paroxysmal dysarthria and ataxia is a rare neurological manifestation of acquired conditions or hereditary disorders, like autosomal dominant episodic dysarthria and truncal ataxia.Pathophysiology of PDA is not fully known to date, and it has been reported in many autoimmune diseases, namely antiphospholipid syndrome and Behcet's disease.Naci Kocer et al., in a study of 65 patients with NBS, demonstrated intra‐axial veins and small vessel pattern on MRI head; commonly, the meso‐diencephalic junction (MDJ) in 46% of patients, followed by ponto‐bulbar region in (40%), the hypothalamic‐thalamic region (23%), basal ganglia in 2%, followed by telencephalon, cerebellum in three, and the cervical cord.Paroxysmal dysarthria and ataxia in neuro‐Behcet's disease, theoretically speaking, is expected if lesions involve the midbrain, as seen in one previously reported case associated with many lesions in the periventricular white matter and brainstem correlating with a typical neuroradiological pattern of NBD fulfilling the NBD diagnostic criteria.Paroxysmal dysarthria and ataxia pathogenesis is not fully understood to date, with other variants as Xia et al. have presented a none‐ataxic PDA variant in solitary midbrain sclerosis. Understanding midbrain PDA triggering lesions, relative to their burden and nature , is paramount, which would aid in better understanding of possible pathophysiology relevant to cerebello‐thalamocortical pathways, and hence better‐targeted treatment.Finally, carbamazepine has been reported to be effective in PDA‐related MS lesions, solitary sclerosis, and APLS‐related lesions, by opposing the ephaptic transmissions between contiguous fibers.The authors have no conflict of interest to declare.AS, LY, and AELB wrote the initial draft of the manuscript. AELB, AELS, and GA contributed to conceptualization and supervision. AS, GA, and A contributed to medical management of the case. AS, LY, AELB, AELS, GA, and A revised the manuscript critically and literature review.This case report was approved by the Hamad Medical Corporation's Medical Research Center (Protocol number: MRC 04‐21‐689).Written informed consent was obtained from the patient for the publication of this case report.


Chromohalobacter canadensis is one of the early halophiles to show promising biotechnology potential, which has not been explored to date. Advanced high throughput technologies such as whole‐genome sequencing allow in‐depth insight into the potential of organisms while at the frontiers of systems biology. At the same time, genome‐scale metabolic models (GEMs) enable phenotype predictions through a mechanistic representation of metabolism. Here, we sequence and analyze the genome of C. canadensis 85B, and we use it to reconstruct a GEM. We then analyze the GEM using flux balance analysis and validate it against literature data on C. canadensis. We show that C. canadensis 85B is a metabolically versatile organism with many features for stress and osmotic adaptation. Pathways to produce ectoine and polyhydroxybutyrates were also predicted. The GEM reveals the ability to grow on several carbon sources in a minimal medium and reproduce osmoadaptation phenotypes. Overall, this study reveals insights from the genome of C. canadensis 85B, providing genomic data and a draft GEM that will serve as the first steps towards a better understanding of its metabolism, for novel applications in industrial biotechnology.Salt tolerant organisms are increasingly being used for the industrial production of high‐value biomolecules due to their better adaptability compared to mesophiles.  Chromohalobacter canadensis 85B was sequenced and its genome‐scale metabolic model was reconstructed and validated, enabling a better understanding of its metabolism. We show that C. canadensis is a metabolically versatile organism with many features for stress and osmotic adaptation, including pathways for compatible solute and polyhydroxybutyrate (PHB) synthesis. The model reveals the ability to grow on several carbon sources in minimal medium, and produce PHBs and compatible solutes such as ectoine, 5‐hydroxyectoine, and betaine.The whole genome of  They can also have a tolerance in the same environment to other conditions such as pH and temperature, thus widening the applications of their bioproducts , MgSO4.7H2O (14 g), KCl (2.8 g), NaBr (0.32 g), NaHCO3 (0.008 g), CaCl2.2H2O (2 g), yeast extract (1 g), Peptone (5 g), and glucose (20 g) as carbon source. The culture was incubated for 3 days at 35°C and 150 rpm in 250 mL Erlenmeyer flasks for polymer production  and visualized on a gel documentation system (BIO‐RAD).From the bacterial cultures, 2 mL of bacterial suspension was obtained for genomic DNA extraction. Genomic DNA was extracted using the PureLink Microbiome DNA purification kit (Invitrogen) according to the manufacturer's instructions. Upon extraction of the pure DNA, an electrophoresis gel was prepared to confirm the presence of a single band corresponding to the whole bacterial genome. A 5 µL of the sample was run on 1% agarose gel for 30 min at 100 v. Gels were stained with ethidium bromide (10 mg mL2.3https://www.patricbrc.org/) using the auto assembly strategy with default parameters  and PATRIC   . Predicted complementary DNA sequences were blasted in the NCBI nonredundant database as well as Swiss‐Prot and UniProt,  and an average G + C content of 60.90%. The N50 length, which is defined as the shortest sequence length at 50% of the genome, was 186,789 bp. The L50 count, which is defined as the smallest number of contigs whose length sum produces N50, was 5  of annotated proteins were included in the subsystems analysis according to the RAST pipeline. An overview of the subsystems for this genome as produced by the annotation pipeline is provided in Figure C. salexigens the first seven groups seem to be the most abundant despite the subtle differences in the relative abundance of orthologous genes between both species. This further emphasizes the importance of these systems in this group of microorganisms.An analysis of orthologous genes shows amino acid metabolism and transport and transcription containing the highest number of orthologous genes Figure . When co3.4C. canadensis 85B and nine subsystems representing biosynthesis and degradation pathways. Predictions show genes for the metabolization of various carbohydrate substrates such as sugar alcohols, C‐1 compounds, sugar acids, monosaccharides, polysaccharides, and fermentation. Enzymes able to metabolize the following substrates were predicted: glucose, starch, sucrose, fructose, mannose, xylose, glycerol, and galactose. The presence of many different pathways for carbohydrate metabolism has significant implications for the adaptation of halophiles.There were 177 carbohydrate metabolism genes in C. salexigens, glucose metabolism occurs exclusively through the Entner–Doudoroff pathway while fructose metabolism occurs through the Entner–Doudoroff and Embden–Meyerhof–Parnas pathways. Fructose metabolism seems to give more metabolic flexibility in response to energy and biosynthetic demands. The Entner–Doudoroff pathway, on the other hand, is inefficient for growth when salinity is low, as a result of metabolite overflow. However, in high salinity, there is a high metabolic burden on this pathway due to the use of NADPH and ATP for the synthesis of compatible solutes. This allows the organism to use other pathways to meet other metabolic requirements  ‐3‐Hydroxybutanoyl‐CoA intermediates that link it to the PHB synthesis pathway.The fatty acid composition of salt‐tolerant organisms is influenced by salt concentrations. This is observed through decreased saturation of fatty acids at suboptimal concentrations. Therefore by varying the ratio of saturated to unsaturated fatty acids adaptation to salt stress can be achieved  is the key enzyme in the PHB synthesis pathway, catalyzing the polymerization of hydroxyalkanoate subunits ‐hydroxyalkanoic acid synthase (C. japonicus). Only one hit was obtained each in the Gene3D, InterPro, Pfam, SUPFAM, and TIGRFAMs, all corresponding to PHA synthase class I. The class I subfamily PHA synthases can polymerize hydroxyacyl‐CoAs with three to five carbons in the hydroxyacyl into PHA esters in this case most likely PHB. These can be accumulated up to 90% of the cell's dry weight. The PhaR genes play a posttranscriptional role and help prevent protease degradation or act directly or indirectly to activate PHA synthase . The annotations are as follows: passive transport (SBO:0000658), active transport (SBO:0000657), cotransport:symport (SBO:0000659), cotransport:antiport (SBO:0000660) other transport reactions (SBO:0000655), general metabolic reactions (SBO:0000176), exchange reactions (SBO:0000627), biomass reactions (SBO:0000629), genes (SBO:0000243), and species (SBO:0000247) , and zinc/iron permease which have been reported to be associated with adaptations to high salt environments by sensing salt stress and regulating intracellular ion homeostasis respectively  showed a decrease in PHB production with increasing nitrogen uptake rates, although with a steeper slope after uptake rates of about 39 mmol/gDW/h , Formal analysis , Funding acquisition , Visualization , Writing – review & editing . Belma Nural Yaman: Conceptualization , Funding acquisition , Writing – review & editing . Chaimaa Tarzi: Formal analysis , Visualization , Writing – review & editing . Pınar Aytar Çelik: Conceptualization , Funding acquisition , Supervision , Writing – review & editing . Mehmet Mutlu: Supervision , Writing – review & editing . Claudio Angione: Conceptualization , Funding acquisition , Supervision , Visualization , Writing – review & editing .None declared.None required.