Search Results (21)

Thaumatin-like proteins (TLPs), a subfamily of pathogenesis-related (PR) proteins, play a vital role in plant defense against pathogens. In this study, 23 CsTLP genes were identified in the Citrus sinensis genome. These genes encode proteins ranging from 203 to 512 amino acids, with molecular weights between 21.88 and 53.75 kDa, classifying them as small molecular weight proteins. The CsTLP genes are unevenly distributed across eight chromosomes, with chromosome 3 containing the highest number (6 genes). Subcellular localization predictions indicate that most CsTLPs are located in the extracellular space. Phylogenetic analysis with Arabidopsis thaliana TLPs classified the CsTLPs into 10 clades, with clade 5 being the largest. Three segmentally duplicated gene pairs were identified, suggesting a mechanism for the expansion of this gene family. Expression profiling revealed tissue-specific patterns, with the highest expression levels observed in roots and leaves. Under biotic stress, qRT-PCR analysis of 12 selected CsTLPs demonstrated pathogen-specific responses: CsTLP9 and CsTLP22 were strongly upregulated during Huanglongbing (HLB, bacterial) infection, by 21.70-fold and 9.47-fold, respectively. Multiple genes, including CsTLP5/13/18/21/23, exhibited over 10-fold upregulation following Citrus Anthracnose (CA, fungal) infection; however, most genes showed only weak responses to Citrus tristeza virus (CTV, viral). These findings underscore the regulatory significance of CsTLPs in pathogen responses and provide an important theoretical foundation for enhancing molecular disease-resistance breeding in Citrus sinensis. Full article

The global citrus industry faces unprecedented challenges due to Huanglongbing (HLB), which is associated with the bacterial pathogen Candidatus Liberibacter asiaticus (CLas). This study evaluates the field performance of 11 rootstocks, grafted with ‘Valencia’ orange (Citrus sinensis), under Florida’s HLB-endemic production conditions, while also examining the impact of systemic applications of oxytetracycline (OTC) via trunk injection. Mature trees received annual OTC injections and were assessed over two production seasons. In year 1, OTC-treated trees exhibited significant improvements regardless of the rootstock, including a 36% increase in yield, an 11% increase in juice TSS, and reduced leaf bacterial titers. During year 2, the positive effects of OTC were sustained, or even enhanced. CLas titers were reduced in both leaves and roots; yield increased by 70%; and fruit weight, juice color, and TSS also improved significantly. Moreover, OTC-injected trees exhibited a larger percentage of finer roots compared to non-injected trees. US-1688 and US-1672, both hybrids of C. maxima ‘Hirado’ and C. reticulata ‘Cleopatra’, emerged as the most productive rootstocks. These results demonstrate the importance of rootstock selection for sustainable citrus cultivation while highlighting the benefits of integrating the systemic delivery of OTC to manage HLB and maximize the resilience of citrus. Full article

Most commercial citrus fruits are grown in acidic soils with high copper (Cu) and low organic matter levels in China. Sweet orange (Citrus sinensis (L.) Osbeck cv. Xuegan) seedlings were treated with 0 (HA0), 0.1 (HA0.1), or 0.5 (HA0.5) mM humic acid (HA) and 0.5 (Cu0.5) or 400 (Cu400 or Cu excess) μM CuCl₂ for 24 weeks. The purpose was to validate the hypothesis that HA reduces the oxidative injury caused by Cu400 in roots and leaves via the coordination of strengthened antioxidant defense and glyoxalase systems. Copper excess increased the superoxide anion production rate by 27.0% and 14.2% in leaves and by 47.9% and 33.9% in roots, the malonaldehyde concentration by 199.6% and 27.8% in leaves and by 369.4% and 77.4% in roots, and the methylglyoxal concentration by 18.2% and 6.6% in leaves and by 381.8% and 153.3% in roots, as well as the H₂O₂ production rate (HPR) by 70.5% and 16.5% in roots, respectively, at HA0 and HA0.5. Also, Cu400 increased the leaf HPR at HA0, but not at HA0.5. The addition of HA reduced the Cu400-induced production and accumulation of reactive oxygen species and methylglyoxal and alleviated the impairment of Cu400 to the antioxidant defense system (ascorbate-glutathione cycle, antioxidant enzymes, sulfur-containing compounds, and sulfur-metabolizing enzymes) and glyoxalase system in roots and leaves. The HA-mediated amelioration of Cu toxicity involved reduced oxidative injury due to the coordination of strengthened antioxidant defense and glyoxalase systems. These findings highlight the promise of HA for sustainable citrus cultivation in heavy metal (Cu)-polluted soils. Full article

Excessive copper (Cu) has become a common physiological disorder restricting the sustainable production of citrus. Coumarin (COU) is a hydroxycinnamic acid that can protect plants from heavy metal toxicity. No data to date are available on the ameliorative effect of COU on plant Cu toxicity. ‘Xuegan’ (Citrus sinensis (L.) Osbeck) seedlings were treated for 24 weeks with nutrient solution containing two Cu levels (0.5 (Cu0.5) and 400 (Cu400) μM CuCl₂) × four COU levels (0 (COU0), 10 (COU10), 50 (COU50), and 100 (COU100) μM COU). There were eight treatments in total. COU supply alleviated Cu400-induced increase in Cu absorption and oxidative injury in roots and leaves, decrease in growth, nutrient uptake, and leaf pigment concentrations and CO₂ assimilation (A_CO2), and photo-inhibitory impairment to the whole photosynthetic electron transport chain (PETC) in leaves, as revealed by chlorophyll a fluorescence (OJIP) transient. Further analysis suggested that the COU-mediated improvement of nutrient status (decreased competition of Cu²⁺ with Mg²⁺ and Fe²⁺, increased uptake of nutrients, and elevated ability to maintain nutrient balance) and mitigation of oxidative damage (decreased formation of reactive oxygen species and efficient detoxification system in leaves and roots) might lower the damage of Cu400 to roots and leaves (chloroplast ultrastructure and PETC), thereby improving the leaf pigment levels, A_CO2, and growth of Cu400-treated seedlings. Full article

Limited data are available on copper (Cu)–pH interaction-responsive genes and/or metabolites in plant roots. Citrus sinensis seedlings were treated with 300 μM (Cu toxicity) or 0.5 μM (control) CuCl₂ at pH 3.0 or 4.8 for 17 weeks. Thereafter, gene expression and metabolite profiles were obtained using RNA-Seq and widely targeted metabolome, respectively. Additionally, several related physiological parameters were measured in roots. The results indicated that elevating the pH decreased the toxic effects of Cu on the abundances of secondary metabolites and primary metabolites in roots. This difference was related to the following several factors: (a) elevating the pH increased the capacity of Cu-toxic roots to maintain Cu homeostasis by reducing Cu uptake and Cu translocation to young leaves; (b) elevating the pH alleviated Cu toxicity-triggered oxidative damage by decreasing reactive oxygen species (ROS) formation and free fatty acid abundances and increasing the ability to detoxify ROS and maintain cell redox homeostasis in roots; and (c) increasing the pH prevented root senescence and cell wall (CW) metabolism impairments caused by Cu toxicity by lowering Cu levels in roots and root CWs, thus improving root growth. There were some differences and similarities in Cu–pH interaction-responsive genes and metabolites between leaves and roots. Full article

Some citrus orchards in China often experience nitrogen (N) deficiency. For the first time, targeted metabolomics was used to examine N-deficient effects on hormones in sweet orange (Citrus sinensis (L.) Osbeck cv. Xuegan) leaves and roots. The purpose was to validate the hypothesis that hormones play a role in N deficiency tolerance by regulating root/shoot dry weight ratio (R/S), root system architecture (RSA), and leaf and root senescence. N deficiency-induced decreases in gibberellins and indole-3-acetic acid (IAA) levels and increases in cis(+)-12-oxophytodienoic acid (OPDA) levels, ethylene production, and salicylic acid (SA) biosynthesis might contribute to reduced growth and accelerated senescence in leaves. The increased ethylene formation in N-deficient leaves might be caused by increased 1-aminocyclopropanecarboxylic acid and OPDA and decreased abscisic acid (ABA). N deficiency increased R/S, altered RSA, and delayed root senescence by lowering cytokinins, jasmonic acid, OPDA, and ABA levels and ethylene and SA biosynthesis, increasing 5-deoxystrigol levels, and maintaining IAA and gibberellin homeostasis. The unchanged IAA concentration in N-deficient roots involved increased leaf-to-root IAA transport. The different responses of leaf and root hormones to N deficiency might be involved in the regulation of R/S, RSA, and leaf and root senescence, thus improving N use efficiency, N remobilization efficiency, and the ability to acquire N, and hence conferring N deficiency tolerance. Full article

Citrus is mainly cultivated in acid soil with low boron (B) and high copper (Cu). In this study, Citrus sinensis seedlings were submitted to 0.5 (control) or 350 μM Cu (Cu excess or Cu exposure) and 2.5, 10, or 25 μM B for 24 weeks. Thereafter, H₂O₂ production rate (HPR), superoxide production rate (SAPR), malondialdehyde, methylglyoxal, and reactive oxygen species (ROS) and methylglyoxal detoxification systems were measured in leaves and roots in order to test the hypothesis that B addition mitigated Cu excess-induced oxidative damage in leaves and roots by reducing the Cu excess-induced formation and accumulation of ROS and MG and by counteracting the impairments of Cu excess on ROS and methylglyoxal detoxification systems. Cu and B treatments displayed an interactive influence on ROS and methylglyoxal formation and their detoxification systems. Cu excess increased the HPR, SAPR, methylglyoxal level, and malondialdehyde level by 10.9% (54.3%), 38.9% (31.4%), 50.3% (24.9%), and 312.4% (585.4%), respectively, in leaves (roots) of 2.5 μM B-treated seedlings, while it only increased the malondialdehyde level by 48.5% (97.8%) in leaves (roots) of 25 μM B-treated seedlings. Additionally, B addition counteracted the impairments of Cu excess on antioxidant enzymes, ascorbate-glutathione cycle, sulfur metabolism-related enzymes, sulfur-containing compounds, and methylglyoxal detoxification system, thereby protecting the leaves and roots of Cu-exposed seedlings against oxidative damage via the coordinated actions of ROS and methylglyoxal removal systems. Our findings corroborated the hypothesis that B addition alleviated Cu excess-induced oxidative damage in leaves and roots by decreasing the Cu excess-induced formation and accumulation of ROS and MG and by lessening the impairments of Cu excess on their detoxification systems. Further analysis indicated that the pathways involved in the B-induced amelioration of oxidative stress caused by Cu excess differed between leaves and roots. Full article

Magnesium is an essential nutrient for the growth and development of plants. Its deficiency is becoming a growing concern in many citrus orchards worldwide, adversely affecting numerous functions in plants and limiting their productivity and quality. Three-year-old orange (Citrus sinensis cv. Newhall) plants grafted on Swingle citrumelo (C. paradisi Macf. × Poncirus trifoliata L.) rootstock were irrigated for 63 days with nutrient solutions containing 0, 12, 24, 48, 96, and 192 mg Mg L⁻¹. Thereafter, Mg deficiency-induced changes were investigated in leaf chlorophyll concentration and fluorescence, and in proline and carbohydrate concentration in leaves and roots, as well as in the nutritional status of leaves, stems, and roots. Magnesium concentration in the nutrient solution was positively correlated with the concentration of Mg (leaves, stems, roots), Ca (rootstock’s stem), K (roots), and Fe (leaves, rootstock’s stem), as well as with the total Mg absorption. However, Mg concentration in the nutrient solution was negatively correlated with the concentration of Mn (rootstock’s stem, roots), Ca (leaves, scion’s stems, roots), and Fe (roots), as well as with the total absorption of Mn, Fe, P, K, and Ca. The lower values of the chlorophyll fluorescence parameters were observed by the effect of the highest concentration of Mg. As far as leaf chlorophyll concentration and carbohydrate and proline content of leaves and roots, they were not affected by the Mg treatments. Concluding, this research highlights the significance of Mg management in citrus farming, offering insights into increasing Mg concentrations, understanding root mechanisms in Mg absorption, and suggesting the benefits of fertilization to address Mg deficiency. It emphasizes the importance of careful Mg fertilizer dosages, considering other nutrient interactions, and provides valuable guidance for optimizing Mg nutrition and overall nutrient management in sweet orange trees. Full article

The effects of copper (Cu)–pH interactions on the levels of hormones and related metabolites (HRMs) in Citrus sinensis leaves and roots were investigated. Our findings indicated that increased pH mitigated Cu toxicity-induced alterations of HRMs, and Cu toxicity increased low-pH-induced alterations of HRMs. Increased pH-mediated decreases in ABA, jasmonates, gibberellins, and cytokinins, increases in (±)strigol and 1-aminocyclopropanecarboxylic acid, and efficient maintenance of salicylates and auxins homeostasis in 300 μM Cu-treated roots (RCu300); as well as efficient maintenance of hormone homeostasis in 300 μM Cu-treated leaves (LCu300) might contribute to improved leaf and root growth. The upregulation of auxins (IAA), cytokinins, gibberellins, ABA, and salicylates in pH 3.0 + 300 μM Cu-treated leaves (P3CL) vs. pH 3.0 + 0.5 μM Cu-treated leaves (P3L) and pH 3.0 + 300 μM Cu-treated roots (P3CR) vs. pH 3.0 + 0.5 μM Cu-treated roots (P3R) might be an adaptive response to Cu toxicity, so as to cope with the increased need for reactive oxygen species and Cu detoxification in LCu300 and RCu300. Increased accumulation of stress-related hormones (jasmonates and ABA) in P3CL vs. P3L and P3CR vs. P3R might reduce photosynthesis and accumulation of dry matter, and trigger leaf and root senescence, thereby inhibiting their growth. Full article

Over-applied copper (Cu)-based agrochemicals are toxic to citrus trees. However, less information is available discussing the ultrastructural alterations in Cu-stressed citrus species. In the present study, seedlings of Citrus sinensis and Citrus grandis that differed in Cu-tolerance were sandy-cultured with nutrient solution containing 0.5 µM Cu (as control) or 300 µM Cu (as Cu toxicity) for 18 weeks. At the end of the treatments, the physiological parameters and ultrastructural features of the citrus leaves and roots were analyzed. The results indicate that Cu toxicity significantly decreased the ratio of shoot biomass to dry weight, the Cu translocation factor and the total chlorophyll of two citrus species. The anatomical and ultrastructural alterations verified that excessive Cu resulted in starch granules accumulated in the leaves and roots of the two citrus species. Under Cu toxicity, increased root flocculent precipitate and thickened root cell wall might reduce the Cu translocation from citrus roots to the shoots. Compared with C. sinensis, C. grandis maintained a relatively integral root cellular structure under Cu toxicity, which provided a structural basis for a higher Cu tolerance than C. sinensis. The present results increase our understanding of the physiological and ultrastructural responses to Cu toxicity in citrus species. Full article

Lasiodiplodia (family Botryosphaeriaceae) is a widely distributed fungal genus that causes a variety of diseases in tropical and subtropical regions. During 2020–2021, a routine survey of fruit tree plants was conducted in five Egyptian Governorates, and fresh samples exhibiting dieback, decline, leaf spot and root rot symptoms were collected. Collection from eight different symptomatic leaves, twigs, branches and roots of fruit trees yielded 18 Lasiodiplodia-like isolates. The sequencing data from the internal transcribed spacer region (ITS), partial translation elongation factor 1-alpha (tef1-a) and β-tubulin (tub2) were used to infer phylogenetic relationships with known Lasiodiplodia species. Two isolates obtained from black necrotic lesions on Phoenix dactylifera leaves were identified as a putative novel species, L. newvalleyensis sp. nov., and were thus subjected to further morphological characterization. The results of isolation and molecular characterization revealed that L. theobromae (n = 9) was the most common species on Mangifera indica, Citrus reticulata, C. sinensis, Ficus carica, Prunus persica, Prunus armeniaca and Pyrus communis trees. Lasiodiplodia pseudotheobromae (n = 5) was isolated from M. indica, Prunus persica and C. sinensis. Lasiodiplodia laeliocattleyae (n = 2) was isolated from C. reticulata. Pathogenicity test results suggested that all Lasiodiplodia species were pathogenic to their hosts. The present study is considered the first to characterize and decipher the diversity of Lasiodiplodia species associated with fruit trees in Egypt, using the multi-locus ITS, tef1-a and tub2 sequence data, along with morphological and pathogenic trials. To our knowledge, this is the first report of L. newvalleyensis on Phoenix dactylifera and L. laeliocattleya on C. reticulata in Egypt and worldwide. Full article

The contribution of reactive oxygen species (ROS) and methylglyoxal (MG) formation and removal in high-pH-mediated alleviation of plant copper (Cu)-toxicity remains to be elucidated. Seedlings of sweet orange (Citrus sinensis) were treated with 0.5 (non-Cu-toxicity) or 300 (Cu-toxicity) μM CuCl₂ × pH 4.8, 4.0, or 3.0 for 17 weeks. Thereafter, superoxide anion production rate; H₂O₂ production rate; the concentrations of MG, malondialdehyde (MDA), and antioxidant metabolites (reduced glutathione, ascorbate, phytochelatins, metallothioneins, total non-protein thiols); and the activities of enzymes (antioxidant enzymes, glyoxalases, and sulfur metabolism-related enzymes) in leaves and roots were determined. High pH mitigated oxidative damage in Cu-toxic leaves and roots, thereby conferring sweet orange Cu tolerance. The alleviation of oxidative damage involved enhanced ability to maintain the balance between ROS and MG formation and removal through the downregulation of ROS and MG formation and the coordinated actions of ROS and MG detoxification systems. Low pH (pH 3.0) impaired the balance between ROS and MG formation and removal, thereby causing oxidative damage in Cu-toxic leaves and roots but not in non-Cu-toxic ones. Cu toxicity and low pH had obvious synergistic impacts on ROS and MG generation and removal in leaves and roots. Additionally, 21 (4) parameters in leaves were positively (negatively) related to the corresponding root parameters, implying that there were some similarities and differences in the responses of ROS and MG metabolisms to Cu–pH interactions between leaves and roots. Full article

Arbuscular mycorrhizal (AM) fungi and endophytic fungi collectively symbiose well with plants and, thus, stimulate plant growth; however, it is not clear whether field inoculation of the fungi enhances the resistance potential of plants, particularly in citrus. In the present study, we inoculated AM fungi (Acaulospora scrobiculata, Diversispora spurca, and D. versiformis) and endophytic fungi (Piriformospora indica) on an eight-year-old lane late navel orange (Citrus sinensis (L.) Osb) trees grafted on Poncirus trifoliata in a field, and we analyzed the response of the leaf antioxidant defense system. Approximately 2 years after inoculation, the root fungal colonization rate and soil hyphal length significantly increased. Fungal inoculation significantly increased the activity of leaf antioxidant enzymes, such as superoxide dismutase, peroxidase, catalase, and ascorbate peroxidase, and the content of non-enzymatic antioxidants, such as reduced ascorbic acid and reduced glutathione. As a result, fungi-inoculated plants maintained lower concentrations of hydrogen peroxide and superoxide anion radicals and lower levels of membrane lipid peroxidation (according to malondialdehyde level) in leaves than uninoculated plants. Among them, inoculation of D. spurca and A. scrobiculata showed relatively higher effects in enhancing the antioxidant defense system than the other fungi. Furthermore, inoculation of D. spurca induced expressions of CsFe-SOD, CsMn-SOD, CsPOD, CsCAT1, and CsPRR7; inoculation of A. scrobiculata and D. versiformis induced expressions of CsCAT1; CsCAT1 and CsPOD were also induced by inoculation of P. indica. All four inoculations almost upregulated expressions of CsFAD6. AM fungi had superior effects than endophytic fungi (e.g., P. indica). According to our findings, inoculation with beneficial fungi, specifically mycorrhizal fungus D. spurca, activated the antioxidant defense system of field citrus trees, thus, having potentially superior resistance in inoculated plants. Full article

Spain is one of the most important producers of lemon fruits in the world (1,250,000 Tm in 2020/2021). In addition, about 80% of the Spanish production of lemons is located in the arid southeast, where fertirrigation is important to ensure the highest productivity. The aim of the present study was to determine the content of nutrients present in two differently grafted lemon trees (Citrus × limon) of the Verna variety, located on a drip-irrigation farm in Librilla (Region of Murcia, Spain). The first one was grafted in a Sweet orange rootstock (Citrus × sinensis), and the second one was grafted in a Bitter orange rootstock (Citrus × aurantium). Both were in 40-year-old trees grown in conventional agricultural practices. The Bitter orange rootstock (Citrus × aurantium) favoured the appearance of the ‘Miriñaque’ (in Spanish) or protuberance of the trunk at the union of the graft, whereas this did not appear in the Sweet orange rootstock (Citrus × sinensis). For the analysis, fresh samples of old leaf, young leaf, and root were collected from five different trees for each rootstock. Once processed after having been weighed, dried, and ground, they were analysed by inductively coupled plasma (ICP) analysis (Optima 3000, PerkinElmer). The result showed that the growth in aerial biomass was higher in Verna grafted on Sweet orange rootstock. The root samples of the lemon tree with Bitter orange rootstock contained a higher amount of Fe, Mn, and Zn than the samples of the lemon tree with Sweet orange rootstock. The rest of the nutrients did not show significant differences. The new and old leaves of Verna in Sweet orange rootstock showed a higher amount of Fe, Mn, and Zn than the new and old leaves of Verna in Bitter orange rootstock. Additionally, the rest of the nutrients did not show significant differences. The study revealed that this protuberance in the trunk prevents the passage of these elements from the root to the aerial part of the trunk. This is probably related to the cell-to-cell passage. Full article

This work compares the tolerance to long-term anoxia conditions (35 days) of five new citrus ‘King’ mandarin (Citrus nobilis L. Lour) × Poncirus trifoliata ((L.) Raf.) hybrids (named 0501XX) and Carrizo citrange (CC, Citrus sinensis (L.) Osb. × Poncirus trifoliata (L.) Raf.), the widely used citrus rootstock in Spain. Growth parameters, chlorophyll concentration, gas exchange and fluorescence parameters, water relations in leaves, abscisic acid (ABA) concentration, and PIP1 and PIP2 gene expressions were assessed. With a waterlogging treatment, the root system biomass of most hybrids went down, and the chlorophyll a and b concentrations substantially dropped. The net CO₂ assimilation rates (An) and stomatal conductance (g_s) lowered significantly due to flooding, and the transpiration rate (E) closely paralleled the changes in g_s. The leaf water and osmotic potentials significantly increased in most 0501 hybrids. As a trend, flooding stress lowered the ABA concentration in roots from most hybrids, but increased in the leaves of CC, 05019 and 050110. Under the control treatment (Ct) conditions, most 0501 hybrids showed higher PIP1 and PIP2 expressions than the control rootstock CC, but were impaired due to the flooding conditions in 05019 and 050110. From this study, we conclude that 0501 genotypes develop some adaptive responses in plants against flooding stress such as (1) stomata closure to prevent water loss likely mediated by ABA levels, and (2) enhanced water and osmotic potentials and the downregulation of those genes regulating aquaporin channels to maintain water relations in plants. Although these traits seemed especially relevant in hybrids 050110 and 050125, further experiments must be done to determine their behavior under field conditions, particularly their influence on commercial varieties and their suitability as flooding-tolerant hybrids for replacing CC, one of the main genotypes that is widely used as a citrus rootstock in Spain, under these conditions. Full article