Sensitive Rootstocks Research Articles

Transcription factors MhDREB2A/MhZAT10 play a role in drought and cold stress response crosstalk in apple.

Drought and cold stresses seriously affect tree growth and fruit yield during apple (Malus domestica) production, with combined stress causing injury such as shoot shriveling. However, the molecular mechanism underlying crosstalk between responses to drought and cold stress remains to be clarified. In this study, we characterized the zinc finger transcription factor ZINC FINGER OF ARABIDOPSIS THALIANA 10 (ZAT10) through comparative analysis of shoot-shriveling tolerance between tolerant and sensitive apple rootstocks. MhZAT10 responded to both drought and cold stresses. Heterologous expression of MhZAT10 in the sensitive rootstock 'G935' from domesticated apple (Malus domestica) promoted shoot-shriveling tolerance, while silencing of MhZAT10 expression in the tolerant rootstock 'SH6' of Malus honanensis reduced stress tolerance. We determined that the apple transcription factor DEHYDRATION RESPONSE ELEMENT-BINDING PROTEIN 2A (DREB2A) is a direct regulator activating the expression of MhZAT10 in response to drought stress. Apple plants overexpressing both MhDREB2A and MhZAT10 genes exhibited enhanced tolerance to drought and cold stress, while plants overexpressing MhDREB2A but with silenced expression of MhZAT10 showed reduced tolerance, suggesting a critical role of MhDREB2A-MhZAT10 in the crosstalk between drought and cold stress responses. We further identified drought-tolerant MhWRKY31 and cold-tolerant MhMYB88 and MhMYB124 as downstream regulatory target genes of MhZAT10. Our findings reveal a MhDREB2A-MhZAT10 module involved in crosstalk between drought and cold stress responses, which may have applications in apple rootstock breeding programs aimed at developing shoot-shriveling tolerance.

PM 7/31 (2) Citrus tristeza virus

<scp>PM</scp> 7/31 (2) Citrus tristeza virus

The tolerance of grapevine rootstocks to copper excess and to the use of calcium and phosphorus to mitigate its phytotoxicity.

High soil copper (Cu) concentrations in vineyards can cause phytotoxicity to grapevine rootstocks. In order to mitigate toxicity, the use of grapevine rootstock genetic variation and the application of amendments are possible strategies. The aim of this study is to assess the tolerance of grapevine rootstocks to Cu excess and whether phosphorus (P) and calcium (Ca) can reduce phytotoxicity caused by Cu. Grapevine rootstock seedlings were produced from selected stakes: Paulsen 1103 (Vitis berlandieri × Vitis rupestris); SO4 (Vitis berlandieri × Vitis riparia); IAC 572 ((Vitis Riparia × Vitis rupestris) × Vitis caribaea); and Isabel (Vitis labrusca). Seedlings were grown in nutrition solution added with the following treatments: 0.3µM Cu (control); 60µM Cu; 60µM Cu and 62mg L-1 P; 60µM Cu and 400mg L-1 Ca. High Cu concentration caused phytotoxicity in all rootstocks, impairing their growth and decreasing nutrient concentration and photosynthetic activity. P and Ca addition had positive effect on the photosynthetic activity of all rootstocks, although it was not enough to revert growth to levels comparable with controls. Overall, based on the results, the application of P and Ca was not efficient in mitigating Cu phytotoxicity in grapevine plants grown in solution. Isabel was the most sensitive rootstock to Cu phytotoxicity, whereas Paulsen 1103 and SO4 presented more tolerance and can be used, together with other management strategies, in contaminated vineyard areas. Therefore, careful genotype rootstock selection for use in high Cu soils is important, while Ca and P are not efficient mitigators of Cu toxicity.

Comparative transcriptomic analyses of citrus cold-resistant vs. sensitive rootstocks might suggest a relevant role of ABA signaling in triggering cold scion adaption

BackgroundThe citrus genus comprises a number of sensitive tropical and subtropical species to cold stress, which limits global citrus distribution to certain latitudes and causes major economic loss. We used RNA-Seq technology to analyze changes in the transcriptome of Valencia delta seedless orange in response to long-term cold stress grafted on two frequently used citrus rootstocks: Carrizo citrange (CAR), considered one of the most cold-tolerant accessions; C. macrophylla (MAC), a very sensitive one. Our objectives were to identify the genetic mechanism that produce the tolerant or sensitive phenotypes in citrus, as well as to gain insights of the rootstock-scion interactions that induce the cold tolerance or sensitivity in the scion.ResultsPlants were kept at 1 ºC for 30 days. Samples were taken at 0, 15 and 30 days. The metabolomic analysis showed a significant increase in the concentration of free sugars and proline, which was higher for the CAR plants. Hormone quantification in roots showed a substantially increased ABA concentration during cold exposure in the CAR roots, which was not observed in MAC. Different approaches were followed to analyze gene expression. During the stress treatment, the 0-15-day comparison yielded the most DEGs. The functional characterization of DEGs showed enrichment in GO terms and KEGG pathways related to abiotic stress responses previously described in plant cold adaption. The DEGs analysis revealed that several key genes promoting cold adaption were up-regulated in the CAR plants, and those repressing it had higher expression levels in the MAC samples.ConclusionsThe metabolomic and transcriptomic study herein performed indicates that the mechanisms activated in plants shortly after cold exposure remain active in the long term. Both the hormone quantification and differential expression analysis suggest that ABA signaling might play a relevant role in promoting the cold hardiness or sensitiveness of Valencia sweet orange grafted onto Carrizo citrange or Macrophylla rootstocks, respectively. Our work provides new insights into the mechanisms by which rootstocks modulate resistance to abiotic stress in the production variety grafted onto them.

Susceptibility of different prunus rootstocks to natural infection of plum pox virus-Turkey (PPV-T) in Central Anatolia

Sharka is the most damaging viral disease of Prunus spp. and the causative agent is plum pox virus (PPV). PPV is widespread and poses a serious problem in stone fruit crops in Turkey. A unique strain of PPV has so far been identified in Turkey and called as PPV-Turkey (PPV-T). The aims of the present study were to describe the efficiency of PPV-T transmission via aphids and to identify the susceptibility of different Prunus rootstocks against PPV-T strain under natural inoculum in Kayseri province located Central Anatolia Region of Turkey. This experiment was established in a small residential orchard where PPV-T infected apricot and plum trees were detected. Five different Prunus rootstocks from which 200 individuals were planted in PPV-T infected orchard and experiment was designed in a randomized complete block desing with 10 replications per block. All rootstocks were regularly tested by DAS-ELISA and RT-PCR during 2015–2017. The results showed that the most susceptible rootstocks in terms of PPV-T infection were Nemaguard, Myrobalan 29B and Myrobalan 29C with the infection rate of 3.61%, 2.74% and 1.04%, respectively. On the other hand, GF677 and Garnem were never found infected with PPV-T. Aphis gossypii, A. craccivora, A. spiraecola, Brachycaudus helichrysi, Hyalopterus pruni, Myzus persicae, Macrosiphum euphorbiae were detected as most abundant aphid species in experimental plot. In 2015, the highest percentage of viruliferous aphids was found as A. gossypii (100%), followed by B. helichrysi (77.8%), A. spiraecola (55.72%), M. persicae (46.05%) and H. pruni (30.76%) whereas it was 100% for A. gossypii and M. euphorbiae followed by A. spiraecola (76.32%), M. persicae (73.1%), H. pruni (62.06%) and B. helichrysi (61.53%) in 2016 by squash real time RT-PCR. The present study showed that PPV-T was effectively transmitted by different aphid species in natural conditions in Kayseri district. It was concluded that Nemaguard and Myrobalan which were found to be the most sensitive rootstocks to PPV-T should not be preferred by growers as Prunus rootstock where PPV poses a high risk.

Prevalence of Mild T30, T3, and Severe VT Strains of &amp;lt;i&amp;gt;Citrus tristeza&amp;lt;/i&amp;gt; Virus in Central-Northern Veracruz, Mexico

The presence and distribution of Citrus tristeza virus (CTV) strains on plants of four citrus species required analysis of plants showing small fruits, yellowing, and twig dieback. Typical citrus tristeza symptoms showed in citrus plantations in Veracruz, Mexico, and caused gradual deterioration, low yield, and death of the plants. Seven citrus-growing municipalities in Central and Northern Veracruz presented the severe VT strain: the incidence in seven of the eight sampled municipalities ranged from 7% in Alamo to 50% in Tihuatlan. As for the T30 strain, its presence and distribution ranged from 17% in Tihuatlan to 83% in Cuitlahuac. Values for the T3 strain went from 20% in Castillo de Teayo to 50% in Papantla. In Tihuatlan, two samples were positive for the T36 strain. Only Citrus sinensis has historical recording infections by severe strains of CTV, unlike other important citrus species. The incidence of orchards quickly declining increases rapidly in Veracruz, the largest citrus producer in Mexico. It becomes critical as the predominant and sensitive rootstock is sour orange (Citrus aurantium). It is important to note that the specific primers used could not be identified in 29% of the samples, suggesting that they could be other than T30, T3, VT, and T36 strains.

Effect of Three Water Regimes on the Physiological and Anatomical Structure of Stem and Leaves of Different Citrus Rootstocks with Distinct Degrees of Tolerance to Drought Stress

Citrus is grown globally throughout the subtropics and semi-arid to humid tropics. Abiotic stresses such as soil water deficit negatively affect plant growth, physiology, biochemistry, and anatomy. Herein, we investigated the effect(s) of three water regimes (control, moderate drought, and severe drought) on the physiological and anatomical structure of 10 different citrus rootstocks with different degrees of tolerance to drought stress. Brazilian sour orange and Gadha dahi performed well by avoiding desiccation and maintaining plant growth, plant water status, and biochemical characters, while Rangpur Poona nucellar (C. limonia) and Sunki × bentake were the most sensitive rootstocks at all stress conditions. At severe water stress, the highest root length (24.33 ± 0.58), shoot length (17.00 ± 1.00), root moisture content (57.67 ± 1.53), shoot moisture content (64.59 ± 1.71), and plant water potential (−1.57 ± 0.03) was observed in tolerant genotype, Brazilian sour orange. Likewise, chlorophyll a (2.70 ± 0.06), chlorophyll b (0.87 ± 0.06) and carotenoids (0.69 ± 0.08) were higher in the same genotype. The lowest H2O2 content (77.00 ± 1.00) and highest proline content (0.51 ± 0.06) were also recorded by Brazilian sour orange. The tolerance mechanism of tolerant genotypes was elucidated by modification in anatomical structures. Stem anatomy at severe drought, 27.5% increase in epidermal cell thickness, 25.4% in vascular bundle length, 30.5% in xylem thickness, 27.7% in the phloem cell area, 8% in the pith cell area, and 43.4% in cortical thickness were also observed in tolerant genotypes. Likewise, leaf anatomy showed an increase of 27.9% in epidermal cell thickness, 11.4% in vascular bundle length, 21% in xylem thickness, and 15% in phloem cell area in tolerant genotypes compared with sensitive ones. These modifications in tolerant genotypes enabled them to maintain steady nutrient transport while reducing the risk of embolisms, increasing water-flow resistance, and constant transport of nutrients across.

Grafting commercial cultivars of almonds on accurate rootstocks mitigates adverse effects of drought stress

Tolerance to drought can be improved by grafting commercial varieties onto drought tolerant rootstocks. The interaction of rootstock and scion might be a mechanism for resistance to the environmental stresses such as drought in almonds. The aim of this research was to study if short-term exposure to drought in grafted plants using diferent rootstocks would show tolerance to drought now. To fulfill this objective, some parameters (proline content, soluble carbohydrates, malondialdehyde (MDA), electrolyte leakage (EL) and antioxidant enzymes activity such as ascorbate peroxidase (ASC) and peroxidase (POD), relative water content (RWC), stem water potential (ΨSWP or SWP) and water use efficiency (WUE)) of scions (5 commercial almonds) grafted on 3 rootstocks were analyzed. Factorial study based on randomized complete block design with three replications was carried out to determine the most tolerance compound. The factors included cultivars in 5 levels (‘Supernova’, ‘Texas’, ‘Marcona’ ‘Shokoufeh’ and ‘K13–40′), rootstocks in 3 levels [(peach × almond hybrids: ‘GF677’, ‘GN22’ and ‘seedling bitter almond ‘No.32′] and drought stress in 4 levels (No: -0.1 MPa (NS), Moderate: -0.7 MPa (MS), Severe: -1.1 MPa (SS) and very severe: ΨSWP ≤ -2.8 MPa (VSS). The results revealed that increasing drought stress leads to a decrease in the proline content, soluble carbohydrates, MDA, EL, and the activity of ASC and POD enzymes in various rootstock-scion grafted were increased. In the VSS, the highest content of proline in comparison with control (+ 73.43%) was related to the ‘K13–40′ on ‘GF677’. The highest MDA (46.50 and 45.29 nmol/gfw) was in ‘Supernova’ and ‘K13–40′ cultivars on the ‘No.32′ rootstock, which increased 76.47% and 75.43%, in the compared to the control, respectively. Drought stress in sensitive rootstock and scion grafted (‘K13–40′ on ‘No.32′), reduced RWC by 44%, also ΨSWP in resistant rootstock and scion grafted (‘Shokoufeh’ on ‘GF677’), WUE increased up to 9 times. In essence, all cultivars, especially ‘Shokoufeh’ grafted on ‘GF677’ rootstock, were more resistant to the VSS in comparison with the grafts of the same cultivars on the ‘No.32′ and ‘GN22’ rootstocks in this study.

Gene Expression under Short-Term Low Temperatures: Preliminary Screening Method to Obtain Tolerant Citrus Rootstocks

Climate change and global warming are leading to a change in weather patterns toward hot and cold waves. Citrus fruits are a tropical or subtropical crop whose growth is altered by changes in weather patterns. Thus, in the present work, two experiments are evaluated to obtain a screening method to select citrus rootstocks that help us to select new low-temperature-tolerant plant materials. One cold experiment was carried out with the Poncirus trifoliata and Citrus macrophylla rootstocks at 4 °C for 4, 8, 24 and 56 h. A second experiment was performed at 4 °C for 5 days with subsequent acclimatization lasting 0, 5, 10 and 24 h. The expression of the cold response genes CAMTA1, CAMTA3, CAMTA5, CBF1, ICE1 and COR413 IM1 was quantified. The results showed that the best rootstock selection strategy was the second experiment, as a higher expression of the genes CAMTA3, CAMTA5, CBF1 and COR413 IM1 was seen in the tolerant genotype P. trifoliata. We quantified the gene expression of proline biosynthesis P5CS1, dOAT and the proline transporters PROT1 and PROT2; the concentration of the amino acid proline in leaves was also quantified. These results once again showed that the best experiment to differentiate between tolerant and sensitive rootstocks was the second experiment with acclimation time.

The Effect of Low Temperatures on Domestic and Wild Pistachio Rootstocks and Interspecific Hybrids Based on Physio-biochemical Indices

In recent decades, spring frost considered a serious threat in pistachio production (Pistacia vera). Considering that, Iran is one of the most important centers of pistachio production and has the highest variety of pistachios in the world, the identification of domestic and wild different rootstocks and their crosses to reach cold-tolerant rootstocks could be one of the effective solutions to face the challenge of spring frost. In this study, cold tolerance was assessed in fourteen rootstocks of pistachio domestic species (Badami Zarand, Sarakhs and Qazvini), interspecific hybrids (Qazvini×Khinjuk, Qazvini×Mutica, Qazvini × Baneh Baghi, Qazvini × Atlantica, Qazvini× Integerrima), non-domesticated species (Atlantica (P. atlantica), Integerrima, Khinjuk, Sarakhs, Mutica (P. atlantica sub. mutica) and Baneh Baghi (P. mutica × P. vera)) and UCB1 hybrid rootstock at low temperatures (4, 0 and -4 °C) for 2 hours. This experiment was conducted in a factorial completely randomized design with three replications on one-year-old seedlings in greenhouse conditions. The results showed that the rate of ion leakage, malondialdehyde and injury index percentage was significant (p <0.01) and their values were decreased in the cold-tolerant rootstocks than the cold-sensitive rootstocks. The interaction between treatments showed that the temperature of - 4 °C caused a significant reduction in ion leakage and malondialdehyde in the cold-tolerant rootstocks. Proline was increased in the cold-tolerant rootstocks compare to cold-sensitive rootstocks. The results of cation leakage percentage were significant in temperature and rootstocks (p <0.01). The results showed that the highest percentage of cation leakage related to potassium and there was a positive and significant correlation between calcium and magnesium and cell membrane stability. According to the results, the most sensitive rootstock was UCB1, Integerrima, Khinjuk and Baneh Baghi, respectively. Hybrids Qazvini , Baneh Baghi, Khinjuk and Integerrima rootstocks were significantly reduced under low-temperature stress. Mutica, Qazvini×Mutica, Sarakhs, Qazvini, Qazvini×Atlantica rootstocks, improved physiological and biochemical traits and maintained cell membrane integrity and finally showed greater tolerance to frost and freezing temperatures, which can be considered in the breeding program.

Stomatal Behaviors and Physiological Responses Affected by Iron Deficiency in Peach Plants Grafted onto Garnem and GF 677

Iron (Fe) is a pivotal nutrient taking roles in respiration, photosynthesis and many plant metabolisms. Chlorosis caused by Fe deficiency generally occurs in high pH environments and many fruit trees including peach are known sensitive to iron starvation. Fe starvation declines leaf chlorophyll and carotenoid contents, supresses plant growth and declines photosynthetic capacity. Iron deficiency induces responses in plants and different stomatal behaviors and physiological alterations occur to sustain the increased iron uptake capacity of Fe-deficient plants. In this study, the effects of iron deficiency on stomatal morphology and leaf physiological responses in peach variety Rich May grafted onto Garnem and GF 677 rootstocks were investigated. Plants were exposed to Fe deficient conditions for 3 months. End of the experiment, many leaf and stomatal properties were evaluated. The highest decrease in plant growth (relative growth rates of shoot diameter and length) induced by Fe deficiency was found in Rich May grafted onto Garnem. SPAD and relative anthocyanin decreased by 52 and 70% in Fe deficient Rich May/Garnem grafting combination. Decreases in SPAD and relative anthocyanin values were lower with GF 677. In GF 677, LRWC decreased by 1.8% and membrane permeability increased by 10%. Fe controlled stomatal behaviors in peach plants. Stomatal factors were found more sensitive to Fe deficiency in sensitive rootstock, Garnem. The increment in stomatal and pore areas leaded increase in stomatal conductance. The damage by Fe depletion was found higher in Garnem due to higher loss in SPAD, anthocyanin and increasing membrane permeability. Thus, GF 677 can be used in peach orchards under Fe deficiency conditions.

Comprehensive analysis of the apple rhizobiome as influenced by different Brassica seed meals and rootstocks in the same soil/plant system

Replant disease refers to the poor growth of trees when attempting to establish the same or related species on old orchard sites. The use of pre-plant Brassicaceae seed meal (SM) soil amendments in combination with apple replant disease-tolerant rootstock genotypes has been shown to be a promising strategy for the control of apple replant disease (ARD). However, optimizing microorganism-driven protection of apple roots from infection by multiple soil-borne pathogens requires a more comprehensive understanding of how “effective” vs. “ineffective” Brassicaceae seed meal × rootstock genotype disease control systems modulate the composition of rhizosphere microbial communities. In particular, the community of oomycetes associated with the apple rhizosphere remains relatively unexplored compared with bacteria and fungi. To address these issues, we sequenced the root associated bacterial, fungal, and oomycete communities of apple replant disease tolerant (G.210) and susceptible (M.26) rootstocks when grown in an orchard replant soil amended with different Brassicaceae seed meal formulations (Brassica juncea + Sinapis alba, B. juncea, and Brassica napus) previously shown to provide varying levels of replant disease control. Multiple microbial components were associated with observed growth differences between “effective” and “ineffective” disease control systems including the absolute abundance of Ilyonectria/Cylindrocarpon in fine root tissue. Amplicon sequencing provided a more detailed picture of the genetic diversity of oomycete groups in the apple rhizosphere than previously appreciated, and highlighted the variability in oomycete community structure between different rootstock × seed meal disease control systems. In Brassica juncea + Sinapis alba SM-structured rhizospheres, the ARD-tolerant rootstock (G.210) harbored higher relative abundances of Peronosporales with reduced potential to infect apple roots and incite replant disease (such as Peronospora destructor and P. acanthicum), whereas the Peronosporales community associated with the sensitive rootstock (M.26) was dominated by the ARD-specific pathogen Phytophthora cactorum. In addition, Brassica juncea + Sinapis alba SM-structured microbiomes were characterized by numerous bacterial and fungal taxa with the potential for biocontrol, biodegradation and bioremediation. Taken together, these results support the hypothesis that particular Brassicaceae SM soil amendments not only provide “effective” disease control, but also promote microbiomes which are likely to contribute to long-term orchard soil health in many other ways. Overall, this comprehensive analysis highlights the significance of the rootstock × seed meal interaction on bacterial, fungal, and oomycete communities within the apple rhizosphere of “effective” vs. “ineffective” disease control systems and the potential influence of these elements on the dynamics of apple replant disease.

Grafting cv. Grechetto Gentile Vines to New M4 Rootstock Improves Leaf Gas Exchange and Water Status as Compared to Commercial 1103P Rootstock

M4 is a relatively new rootstock that was selected for increased resilience of vineyards across hot regions where meteorological drought is often coupled to water scarcity. However, M4 has thus far been tested only against water-stress sensitive rootstocks. Against this backdrop, the aim of the present work is to examine the water status and gas exchange performances of vines grafted to M4 in comparison to those of vines grafted to a commercial stock that is drought-tolerant, 1103 Paulsen (1103P), under a progressive water deficit followed by re-watering. This study was undertaken on Grechetto Gentile, a cultivar that is renowned for its rather conservative water use (near-isohydric behavior). While fifty percent of both grafts were fully irrigated (WW), the remaining underwent progressive water stress by means of suspending irrigation (WS). Soil and leaf water status, as well as leaf gas exchanges, along with chlorophyll fluorescence, were followed daily from 1 day pre-stress (DOY 176) until re-watering (DOY 184). Final leaf area per vine, divided in main and lateral contribution, was also assessed. While 1103P grafted vines manifested higher water use under WW conditions, progressive stress evidenced a faster water depletion by 1103P, which also maintained slightly more negative midday leaf water potential (Ψleaf) as compared to M4 grafted plants. Daily gas exchange readings, as well as diurnal assessment performed at the peak of stress (DOY 183), also showed increased leaf assimilation rates (A) and water use efficiency (WUE) in vines grafted on M4, which were also less susceptible to photosynthetic downregulation. Dynamic of stomatal closure targeted at 90% reduction of leaf stomatal conductance showed a similar behavior among rootstocks since the above threshold was reached by both at Ψleaf of about −1.11 MPa. The same fractional reduction in leaf A was reached by vines grafted on M4 at a Ψleaf of −1.28 MPa vs. −1.10 MPa measured in 1103P, meaning that using M4 as a rootstock will postpone full stomatal closure. While mechanisms involved in improved CO2 uptake in M4-grafted vines under moderate-to-severe stress are still unclear, our data support the hypothesis that M4 might outscore the performance of a commercial drought-tolerant genotype (1103P) and can be profitably used as a tool to improve the resilience of vines to summer drought.

Tolerance of citrus rootstocks to poor water quality is improved by root zone aeration via selective uptake of ions, higher photosynthesis and carbon storage

The effects of root zone aeration and irrigation water quality (WQ) were studied in two rootstocks with contrasting tolerances to abiotic stress. We hypothesized that the main influence of TWW is that of its salinity (especially Na and K and their ratios), that the organic load and its BOD have a marginal influence and that aeration facilitates selectivity of roots and/or leaves and therefore improves plant performance. The factorial experiment included root zone aeration, fresh water (FW), TWW, and fresh water with a salt concentration similar to TWW (FW + NaCl). One year old unworked rootstocks of sensitive Citrus volkameriana (Volka) and tolerant X639 (Cleopatra mandarin X Poncirus trifoliata) were treated for 4 months, followed by 2 months of recovery. Measurements included ionic and water relations, photosynthetic parameters, growth, morphology, tissue specific membrane properties, and sugar and starch contents. Volka was highly sensitive to WQ and aeration while X639 was influenced much less. During the first two months aeration increased Volka leaf photosynthesis and gas exchange by 15%, water and potassium (K) uptake by 20%, carbon storage by 15% and biomass by 20%; which compensated for the negative effects of poor WQ. Changes in distribution of K and Na ions hint at the mechanisms for the aeration influence on tolerance to poor WQ. Results indicate that aeration enhanced K transport from the roots to leaves and reduced Na concentrations in the roots of the sensitive rootstock. These, as well as changes in the wide range of leaf and root parameters measured, are discussed with relation to their implications for our understanding of plant response and tolerance to low WQ in poorly drained soils.

Rootstock-induced molecular responses associated with drought tolerance in sweet orange as revealed by RNA-Seq

BackgroundCitrus plants are commercially propagated by grafting, with the rootstock variety influencing a number of horticultural traits, including drought tolerance. Among the different rootstock varieties available for citrus propagation, ‘Rangpur’ lime is known to confer enhanced tolerance to drought as compared to other citrus rootstocks. The objective of this study was to investigate the poorly understood molecular responses underlying the rootstock-induced drought tolerance in sweet orange.ResultsRNA-Seq transcriptome analysis was carried out in leaves of sweet orange grafted on ‘Rangpur’ lime subjected to control and drought-stress treatments, under greenhouse conditions, using the Illumina HiSeq platform. A total of 41,827 unique transcripts were identified, among which 1764 transcripts showed significant variation (P ≤ 0.001) between the treatments, with 1081 genes induced and 683 repressed by drought-stress treatment. The transcripts were distributed in 44 different categories of cellular component, molecular function and biological process. Several genes related to cell metabolism, including those involved in the metabolisms of cell wall, carbohydrates and antioxidants, light reactions, biotic and abiotic stress responses, as well as genes coding for transcription factors (TFs), protein kinases (PKs) and proteins involved in the abscisic acid (ABA) and ethylene signaling pathways, were differentially regulated by drought stress. RNA-Seq data were validated by quantitative real-time PCR (qPCR) analysis and comparative analysis of expression of the selected genes between sweet orange grafted on drought-tolerant and -sensitive rootstocks revealed new candidate genes for drought tolerance in citrus.ConclusionsIn conclusion, our results showed that only a relatively small but functionally diverse fraction of the sweet orange transcriptome, with functions in metabolism, cellular responses and regulation, was differentially regulated by drought stress. The data suggest that the rootstock-induced drought tolerance in sweet orange includes the transcriptional activation of genes related to the cell wall, soluble carbohydrate and antioxidant metabolisms, biotic and abiotic stress responses, TFs, PKs and ABA signaling pathway, and the downregulation of genes involved in the starch metabolism, light reactions and ethylene signaling. Future efforts to elucidate their functional roles and explore their potential in the citrus genetic improvement should benefit from this data.

Regulated deficit irrigation benefits the production of container-grown citrus nursery trees

Withholding irrigation at − 15 to − 25 kPa water potential significantly saved water with minimal changes in plant growth, water relations, and percentage of shippable citrus trees on drought-tolerant and -sensitive rootstocks. We evaluated the impacts of regulated deficit irrigation (RDI) on plant growth, water relations, and biochemical variables of Valencia sweet orange nursery trees grafted on drought-tolerant Rangpur lime (RL) and drought-sensitive Swingle citrumelo (SC) rootstocks in pots. Irrigation was withheld to potting mix water potential of − 15 (mild RDI) and − 25 kPa (moderate RDI), with daily irrigation as control, in three graft growth periods (20–60, 61–120, and 20–120 days after grafting, DAG). Plant growth was reduced by moderate RDI, albeit RL induced more vigor than SC. SC induced higher CO2 assimilation rates and water use efficiency (WUE), regardless of the irrigation regime, and only trees grafted on RL decreased WUE under moderate RDI. Most leaf nutrient concentrations were increased with RDI intensity, while K levels were decreased by moderate RDI. Starch reserves were drastically decreased, while reducing sugars were increased by RDI in all plant organs, most notably in the leaves; roots represented the main source of carbohydrates for both rootstocks under water deficit. Starch concentration promptly recovered in all organs after restoring irrigation. Leaf proline concentration was 20 times higher at RDI, but decreased by 50% just 1 day after rehydration. Mild-to-moderate RDI resulted in water savings of 54–80%, but a 0–40% reduction in the percentage of shippable trees at 120 DAG; the biological responses were mainly related to the RDI duration. The production of container-grown citrus nursery trees in greenhouses could greatly benefit from the use of RDI, without major negative effects on tree quality.

Drought tolerance in citrus rootstocks is associated with better antioxidant defense mechanism

Citrus is one of the major fruit crops grown worldwide. Citrus trees are affected by different abiotic stresses including drought which decrease its yield. In this study, six different citrus rootstocks (Volkameriana lemon, Brazilian sour orange, Carrizo citrange, Eureka lemon, Gada dahi, and Rangpur lime) were subjected to drought stress (24 days) and leaf relative water content (LRWC), chlorophyll 'a' and 'b', antioxidant capacity (AC), total phenolic content, and proline content (PRO) were measured. Hydrogen peroxide (H2O2), lipid peroxidation (MDA), total soluble proteins (TSP), and enzymatic antioxidant activities, such as superoxidase dismutase (SOD), catalase (CAT), and peroxidase (POD), were measured in leaves and roots of the rootstocks. Drought-stressed Volkameriana lemon had the minimum LRWC but higher SOD activity in its leaves. Carrizo citrange possessed the minimum amount of Chl 'a' and 'b' in leaves and exhibited lesser MDA and H2O2 in roots but greater TSP content in leaves and roots. SOD, CAT, POD, PRO, MDA, H2O2, and AC decreased in the leaves of Eureka lemon and Rangpur lime under drought stress. MDA and H2O2 contents were observed higher in leaves of Brazilian sour orange (a drought sensitive rootstock). However, higher SOD activity in roots and higher CAT and POD activities in leaves and roots were recorded in Carrizo citrange (a drought-tolerant rootstock). The results indicated that all the studied rootstocks were different in their defense mechanism. Tolerant rootstocks exhibited less amount of MDA and H2O2 and more antioxidant enzymatic activities (SOD, CAT, and POD) to cope with reactive oxygen species produced during drought stress.

Polyethylene glycol damages grafted citrus plants based on biometric, physiological,and biochemical responses

Abstract Drought tolerance is a major trait for the selection of citrus rootstocks in breeding programs. PEG 4000 solution at 110 g L-1 (osmotic potential of -376 kPa) was used to evaluate biometric, physiological, and biochemical responses of grafted citrus plants in pots. ‘Valencia’ sweet orange plants grafted onto ‘Rangpur’ lime (drought tolerant) rootstock showed greater plant growth compared to that on ‘Swingle’ citrumelo (drought sensitive) rootstock, regardless of PEG-induced stress; however, biological responses to PEG were similar on both rootstocks. Plant dry weight, gas exchange, and leaf and root starch concentrations were reduced by PEG. Water potential at 2:00 p.m. and Fe and Mn leaf concentrations increased with severe phytotoxic symptoms. The results corroborate the limitation of PEG for assessing drought tolerance of citrus plants at the whole plant level.

Characterization of Citrus WRKY transcription factors and their responses to phytohormones and abiotic stresses

WRKY transcription factors (TF) family is involved in a huge variety of plant processes, including seed germination, plant development, phytohormone signalling, and defence against both biotic or abiotic stresses. In this work, WRKY TF family has been characterized in citrus. In a first experiment, the relative expression of CsWRKYs was analyzed in shoots and roots of plants treated with abscisic acid (ABA), salicylic acid (SA), and methyl jasmonate (MeJA) under in vitro conditions. Expression of CsWRKYs was also determined in roots of commercial citrus rootstocks subjected to osmotic and salt stresses. A total amount of 50 CsWRKYs has been found and devided into different groups of WRKY family according to the WRKY domain sequences. In response to the applications of phytohormones, the highest differences were observed in roots, and it was found that ABA and SA generally repressed CsWRKYs expressions, but MeJA induced their overexpression. Osmotic stress repressed the expression of most of the CsWRKYs, whereas salt stress induced their expression. Moreover, salt stress induced higher increase in CsWRKY expressions in the tolerant rootstock Citrus macrophylla than in the sensitive rootstock Carrizo citrange, suggesting that these TFs may play an important role in response to this stress.

Changes in Morphological, Physiological Traits and Enzyme Activity of Grafted and Ungrafted Grapevine Rootstocks Under Drought Stress

American grapevine rootstocks are used as the most efficient control method for phylloxera disease. Vitis vinifera species are grown over these rootstocks. The rootstocks may have different drought tolerances. In present study, morphological and physiological responds and enzyme activities of grafted and ungrafted grapevines were investigated under drought stress. The rootstocks Rupestris du Lot, 420A, 5BB, SO4, 8B, 110R, 1103P, 140Ruggeri and Ramsey and K‑7 clone of Sultani Seedless grape cultivar were used as the plant material of this study. Drought stress was exerted as water deficits and stress treatments were terminated when the available moisture level dropped to 5%. Current findings revealed that grafted plants were less influenced morphologically from the droughts than the ungrafted ones. In control group, average leaf water potential was measured as −0.51 MPa for ungrafted plants and −0.46 MPa for grafted plants. In stress group, leaf water potential was measured as −2.03 MPa for ungrafted plants and as −1.69 MPa for grafted ones. Superoxide dismutase (SOD) and ascorbate peroxidase (APX) enzyme activities of both grafted and ungrafted plants increased with drought stress. Considering the enzyme activities of ungrafted plants in control and stress groups, it was observed that SOD activity increased from 508 (unit/mg protein) to 852 (unit/mg protein); APX activity increased from 153 (micromole/mg protein/min) to 584 (micromole/mg protein/min). In grafted plants, SOD activity increased from 639 (unit/mg protein) to 796 (unit/mg protein); APX activity increased from 414 (micromole/mg protein/min) to 1002 (micromole/mg protein/min). The drought resistant rootstocks (like 140Ru, 110R) exhibited higher enzyme activity in both control and stress groups than drought sensitive rootstocks (like 5BB, SO4).