Insight into Physiological and Biochemical Determinants of Salt Stress Tolerance in Tetraploid Citrus.

Citrus are classified as salt-sensitive crops. However, a large diversity has been observed regarding the trends of tolerance among citrus. In the present article, physiological and biochemical studies of salt stress tolerance were carried out according to the level of polyploidy of different citrus genotypes. We particularly investigated the impact of tetraploidy in trifoliate orange (Poncirus trifoliata (L.) Raf.) (PO4x) and Cleopatra mandarin (Citrus reshni Hort. Ex Tan.) (CL4x) on the tolerance to salt stress compared to their respective diploids (PO2x and CL2x). Physiological parameters such as gas exchange, ions contents in leaves and roots were analyzed. Roots and leaves samples were collected to measure polyphenol, malondialdehyde (MDA), ascorbate and H2O2 contents but also to measure the activities of enzymes involved in the detoxification of active oxygen species (ROS). Under control conditions, the interaction between genotype and ploidy allowed to discriminate different behavior in terms of photosynthetic and antioxidant capacities. These results were significantly altered when salt stress was applied when salt stress was applied. Contrary to the most sensitive genotype, that is to say the diploid trifoliate orange PO2x, PO4x was able to maintain photosynthetic activity under salt stress and had better antioxidant capacities. The same observation was made regarding the CL4x genotype known to be more tolerant to salt stress. Our results showed that tetraploidy may be a factor that could enhance salt stress tolerance in citrus.

GABA shunt pathway is stimulated in response to early defoliation-induced carbohydrate limitation in Mandarin fruits.

The regulation of sugar and organic acid metabolism during fruit development has a major effect on high-quality fruit production. The reduction of leaf area is a common feature in plant growth, induced by abiotic and biotic stresses and disturbing source/sink ratio, thus impacting fruit quality. Here, we induced carbohydrate limitation by partial leaf defoliation at the beginning of the second stage of mandarin development (before the citrate peak). Resulting changes were monitored in the short-term (48 h and 1 week) and long-term (7 weeks) after the defoliation. Short-term response to early defoliation implied metabolic settings to re-feed TCA for sustaining respiration rate. These features involved (i) vacuolar sucrose degradation (high acid invertase activity and mRNA expression level) and enhanced glycolytic flux (high ATP-phosphofructokinase activity), (ii) malic and citric acid utilization (increased phosphoenolpyruvate kinase and NADP-Isocitrate dehydrogenase) associated with vacuolar citric acid release (high mRNA expression of the transporter CsCit1) and (iii) stimulation of GABA shunt pathway (low GABA content and increased mRNA expression of succinate semialdehyde dehydrogenase). A steady-state proline level was found in ED fruits although an increase in P5CS mRNA expression level. These results contribute to a better knowledge of the molecular basis of the relationship between defoliation and sugar and organic acid metabolism in mandarin fruit.

Enhanced Photosynthetic Capacity, Osmotic Adjustment and Antioxidant Defenses Contribute to Improve Tolerance to Moderate Water Deficit and Recovery of Triploid Citrus Genotypes.

Currently, drought stress is a major issue for crop productivity, and future climate models predict a rise in frequency and severity of drought episodes. Polyploidy has been related to improved tolerance of plants to environmental stresses. In Citrus breeding programs, the use of triploidy is an effective way to produce seedless fruits, one of the greatest consumer expectations. The current study used physiological and biochemical parameters to assess the differential responses to moderate water deficit of 3x genotypes compared to 2x genotypes belonging to the same hybridization. Both parents, the mandarin Fortune and Ellendale tangor, were also included in the experimental design, while the 2x common clementine tree was used as reference. Water deficit affects leaf water status, as well as physiological and detoxification processes. Triploid genotypes showed a better ability to maintain water status through increased proline content and photosynthetic capacity. Moreover, less oxidative damage was associated with stronger antioxidant defenses in triploid genotypes. We also found that triploidy improved the recovery capacity after a water deficit episode.

Improved response of triploid citrus varieties to water deficit is related to anatomical and cytological properties.

Polyploidy plays a major role in citrus plant breeding to improve the adaptation of polyploid rootstocks as well as scions to adverse conditions and to enhance agronomic characteristics. In Citrus breeding programs, triploidy could be a useful tool to react to environmental issues and consumer demands because the produced fruits are seedless. In this study, we compared the physiological, biochemical, morphological, and ultrastructural responses to water deficit of triploid and diploid citrus varieties obtained from 'Fortune' mandarin and 'Ellendale' tangor hybridization. One diploid clementine tree was included and used as a reference. All studied scions were grafted on C-35 citrange rootstock. Triploidy decreased stomatal density and increased stomata size. The number of chloroplasts increased in 3x varieties. These cytological properties may explain the greater photosynthetic capacity (Pnet, gs, Fv/Fm) and enhanced water-holding capacity (RWC, proline). In addition, reduced degradation of ultrastructural organelles (chloroplasts and mitochondria) and thylakoids accompanied by less photosynthetic activity and low oxidative damages were found in 3x varieties. Triploid varieties, especially T40-3x, had a better ability to limit water loss and dissipate excess energy (NPQ) to protect photosystems. Higher starch reserves in 3x varieties suggest a better carbon and energy supply and increases in plastoglobuli size suggest less oxidative damage (H2O2, MDA), especially in T40-3x, and preservation of photosynthetic apparatus. Taken together, our results suggest that desirable cytological and ultrastructural traits induced by triploidy improve water stress response and could be a useful stress marker during environmental constraints.

Triploid Citrus Genotypes Have a Better Tolerance to Natural Chilling Conditions of Photosynthetic Capacities and Specific Leaf Volatile Organic Compounds.

Low temperatures during winter are one of the main constraints for citrus crop. Polyploid rootstocks can be used for improving tolerance to abiotic stresses, such as cold stress. Because the produced fruit are seedless, using triploid scions is one of the most promising approaches to satisfy consumer expectations. In this study, we evaluated how the triploidy of new citrus varieties influences their sensitivity to natural chilling temperatures. We compared their behavior to that of diploid citrus, their parents (Fortune mandarin and Ellendale tangor), and one diploid clementine tree, as reference, focusing on photosynthesis parameters, oxidative metabolism, and volatile organic compounds (VOC) in leaves. Triploid varieties appeared to be more tolerant than diploid ones to natural low temperatures, as evidenced by better photosynthetic properties (Pnet, gs, Fv/Fm , ETR/P net ratio), without relying on a better antioxidant system. The VOC levels were not influenced by chilling temperatures; however, they were affected by the ploidy level and atypical chemotypes were found in triploid varieties, with the highest proportions of E-ß-ocimene and linalool. Such compounds may contribute to better stress adaptation.

Triploidy in Citrus Genotypes Improves Leaf Gas Exchange and Antioxidant Recovery From Water Deficit.

The triploidy has proved to be a powerful approach breeding programs, especially in Citrus since seedlessness is one of the main consumer expectations. Citrus plants face numerous abiotic stresses including water deficit, which negatively impact growth and crop yield. In this study, we evaluated the physiological and biochemical responses to water deficit and recovery capacity of new triploid hybrids, in comparison with diploid hybrids, their parents ("Fortune" mandarin and "Ellendale" tangor) and one clementine tree used as reference. The water deficit significantly decreased the relative water content (RWC) and leaf gas exchange (P net and g s ) and it increased the levels of oxidative markers (H2O2 and MDA) and antioxidants. Compared to diploid varieties, triploid hybrids limited water loss by osmotic adjustment as reflected by higher RWC, intrinsic water use efficiency (iWUE Pnet/gs ) iWUE and leaf proline levels. These had been associated with an effective thermal dissipation of excess energy (NPQ) and lower oxidative damage. Our results showed that triploidy in citrus enhances the recovery capacity after a water deficit in comparison with diploids due to better carboxylation efficiency, restored water-related parameters and efficient antioxidant system.

Effect of chronic arsenic exposure under environmental conditions on bioaccumulation, oxidative stress, and antioxidant enzymatic defenses in wild trout Salmo trutta (Pisces, Teleostei).

The present study evaluates the relation between chronic arsenic (As) exposure in the natural distribution area of wild brown trout (Salmo trutta), oxidative stress and antioxidant enzymatic defenses. Two rivers of the same watershed were evaluated to highlight the correlation between As accumulation and the resulting stress: (i) the Presa River, which has high chronic As levels (2281.66 µg/L) due to past mining activity, and (ii) the Bravona River (control river). This metalloid was measured in main fish tissues (gills, kidney, liver, muscle, gonads and fins) and water. As organotropism in S. trutta was kidney > liver > gill > fin > gonad > muscle. The HepatoSomatic Index (HSI) and somatic condition (CF) were used to compare fish population conditions from both sites. Arsenic can be absorbed by the gills and can induce oxidative stress and disturb antioxidant defenses. The aim of this study was to evaluate oxidative stress response by measuring malondialdehyde (MDA) content, as a marker of lipid peroxidation, and antioxidant enzymatic defenses (Superoxide dismutase (SOD), catalase CAT, glutathione peroxidase (GPx) and glutathione S-transferase (GST)), in the main tissues of control and exposed trout. The highest MDA content was found in the kidney and liver of exposed trout. SOD and CAT activities in exposed livers and kidneys were considerably increased while a significant rise of GPx activity was observed only in the liver. GST activity was found to be significantly induced in the liver of exposed trout. The results demonstrate that arsenic bioaccumulation can induce lipid peroxidation and substantial modifications in antioxidant enzymatic defenses in main wild trout tissues.

Tetraploid Carrizo citrange rootstock (Citrus sinensis Osb.×Poncirus trifoliata L. Raf.) enhances natural chilling stress tolerance of common clementine (Citrus clementina Hort. ex Tan).

Low temperatures can disturb the development, growth and geographic distribution of plants, particularly cold-sensitive plants in the Mediterranean area, where temperatures can reach seasonally low levels. In citrus crops, scion/rootstock combinations are used to improve fruit production and quality, and increase tolerance to biotic and abiotic stresses. In the last decade, several studies have shown that tetraploid citrus seedlings or rootstocks are more tolerant to abiotic stress than their respective diploid. The objective of this study was to test whether the use of tetraploid rootstocks can improve the chilling tolerance of the scion. We compared physiological and biochemical responses to low seasonal temperatures of common Clementine (Citrus sinensis Osb.×Poncirus trifoliata L. Raf.) grafted on diploid and tetraploid Carrizo citrange rootstocks, named C/2xCC and C/4xCC, respectively. During the coldest months, C/4xCC showed a smaller decrease in net photosynthesis (Pn), stomatal conductance (Gs), chlorophyll fluorescence (Fv/Fm), and starch levels, and lower levels of malondialdehyde and electrolyte leakage than C/2xCC. Specific activities of catalase (CAT), ascorbate peroxidase (APX) and dehydroascorbate reductase (DHAR) were higher in C/4xCC during the cold period, whereas chlorophyll, proline, ascorbate and hydrogen peroxide (H2O2) levels and superoxide dismutase (SOD) activity did not vary significantly between C/4xCC and C/2xCC throughout the study period. Taken together, these results demonstrate that tetraploid Carrizo citrange rootstock improves the chilling tolerance of common clementine (scion) thanks to a part of the antioxidant system.